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Diagnostic accuracy of serological diagnosis of hepatitis C and B using dried blood spot samples (DBS): two systematic reviews and meta-analyses

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BMC Infectious DiseasesBMC series – open, inclusive and trusted201717 (Suppl 1) :700

https://doi.org/10.1186/s12879-017-2777-y

  • Published:

Abstract

Background

Dried blood spots (DBS) are a convenient tool to enable diagnostic testing for viral diseases due to transport, handling and logistical advantages over conventional venous blood sampling. A better understanding of the performance of serological testing for hepatitis C (HCV) and hepatitis B virus (HBV) from DBS is important to enable more widespread use of this sampling approach in resource limited settings, and to inform the 2017 World Health Organization (WHO) guidance on testing for HBV/HCV.

Methods

We conducted two systematic reviews and meta-analyses on the diagnostic accuracy of HCV antibody (HCV-Ab) and HBV surface antigen (HBsAg) from DBS samples compared to venous blood samples. MEDLINE, EMBASE, Global Health and Cochrane library were searched for studies that assessed diagnostic accuracy with DBS and agreement between DBS and venous sampling. Heterogeneity of results was assessed and where possible a pooled analysis of sensitivity and specificity was performed using a bivariate analysis with maximum likelihood estimate and 95% confidence intervals (95%CI). We conducted a narrative review on the impact of varying storage conditions or limits of detection in subsets of samples. The QUADAS-2 tool was used to assess risk of bias.

Results

For the diagnostic accuracy of HBsAg from DBS compared to venous blood, 19 studies were included in a quantitative meta-analysis, and 23 in a narrative review. Pooled sensitivity and specificity were 98% (95%CI:95%–99%) and 100% (95%CI:99–100%), respectively. For the diagnostic accuracy of HCV-Ab from DBS, 19 studies were included in a pooled quantitative meta-analysis, and 23 studies were included in a narrative review. Pooled estimates of sensitivity and specificity were 98% (CI95%:95–99) and 99% (CI95%:98–100), respectively. Overall quality of studies and heterogeneity were rated as moderate in both systematic reviews.

Conclusion

HCV-Ab and HBsAg testing using DBS compared to venous blood sampling was associated with excellent diagnostic accuracy. However, generalizability is limited as no uniform protocol was applied and most studies did not use fresh samples. Future studies on diagnostic accuracy should include an assessment of impact of environmental conditions common in low resource field settings. Manufacturers also need to formally validate their assays for DBS for use with their commercial assays.

Background

It is estimated that 71 million people are infected with hepatitis C (HCV) (defined as those with viraemic infection) and 257 million with hepatitis B (HBV) (defined as HBsAg positive) worldwide [14]. Both HBV and HCV infection predominantly affect persons in low and middle income countries [2, 5]. Late complications of HBV and HCV infection are cirrhosis and hepatocellular carcinoma. Overall, there were 1.34 million deaths attributable to these complications in 2015 with a trend towards increasing deaths since 1990 [2]. Most patients are not aware of their infection until they have advanced complications of the disease [6] due to lack of access to and affordability of testing. Just as an increase in access to treatment is important [7, 8] early identification is paramount and also cost-effective [9] to avoid disease complications.

HBsAg and HCV-Ab screening is traditionally done by serology using either laboratory based enzyme-immuno-assay (EIA) or a point-of-care rapid diagnostic test (RDT). While several rapid lateral flow tests exist for HBsAg and HCV-Ab detection, their quality is variable or unknown and in particular there is a paucity of good quality HBsAg RDTs available on the market [10, 11]. Four HCV-Ab RDTs are WHO prequalified as of August 2017 but none of the HBsAg RDTs have met the requirements for WHO prequalification [12]. The choice of which format of serological assays to use in a programmatic setting will depend on a variety of factors; most importantly, ease of use and the characteristics of the testing site, such as storage facilities, infrastructure, level of staff skills and cost. Confirmation of active viraemic infection is done using nucleic acid testing (NAT) for HCV RNA and HBV DNA or using serological assays for detection of HCV core antigen.

In order to facilitate more widespread uptake of testing for HBV and HCV, there needs to be greater access to diagnostic assays. The use of dried blood spots (DBS) for transportation using fingerstick (in adults and older children), or heel pricking (in neonates and infants) sampling of capillary blood and subsequent analysis with automated high-throughput laboratory-based EIA represents another affordable alternative to testing using RDTs, particularly in settings with limited infrastructure. Another advantage of DBS use in low resource settings [13] is that capillary blood collection does not require a trained health worker to perform venipuncture and that DBS utilizing capillary blood needs less volume than venepuncture. Furthermore, no centrifuge or even basic laboratory facilities with electrical power are needed to prepare plasma [14]; and since transport and handling do not require high skill or a cold chain, the risk of contamination is reduced [15]. The main disadvantage of DBS is that the existing commercial assays have not been validated or received regulatory approval with this method of sample collection and transport.

Despite this limitation, DBS have been increasingly used in recent years to screen for a number of viral diseases, including HIV and viral hepatitis [13, 1619]. Some studies have suggested that the use of DBS may increase uptake of hepatitis testing among certain vulnerable risk groups [2022]. While there have been several systematic reviews on diagnostic performance of RDTs for HBsAg and HCV-Ab [10, 11, 23] and on the use of POC tests in viral hepatitis testing [10, 24, 25], and various validation studies of diagnostic accuracy studies aiming to validate DBS have been performed [26, 27] including a systematic review of HCV RNA detection with DBS [28], to our knowledge there has been no summary of evidence on diagnostic accuracy for HBsAg and HCV-Ab testing using DBS. We have conducted two systematic reviews and meta-analyses: one on the diagnostic accuracy of HCV-Ab and the other on the diagnostic accuracy of HBsAg from DBS samples compared to venous samples in persons identified for hepatitis testing. This review informed the WHO guidelines on testing for chronic HBC and HCV in low and middle income countries [29] and was conducted in conjunction with systematic reviews on the diagnostic accuracy of the detection of HBV DNA and HCV RNA from DBS [30].

Methods

Search strategy and selection criteria

PRISMA reporting guidelines were followed [31] and the QUADAS-2 tool was used to estimate quality of studies as a risk of bias tool [32]. We conducted two systematic reviews and meta-analyses on the diagnostic accuracy of HBsAg and HCV antibody detection from DBS compared to venous blood. We searched English language studies using five databases (MEDLINE, Web of Science, EMBASE, Global Health and LILACS) with the following search terms: DBS, Dried blood spot, Dry blood spot, filter paper, Guthrie paper, hepatitis, hepatitis B, hepatitis C, HBsAg, HBV, HCV, HCV RNA, HBV DNA (adapted to databases) on 1.9.2015 and updated it on 22.8.2017.

Abstracts were included for fulltext review if inclusion criteria were fulfilled (namely DBS samples and plasma or serum samples used for detection of HCV-Ab and/or HBsAg) or if exclusion of the abstract could not be performed solely on the basis of the information of the abstract. Eligible studies included comparisons of an index test HCV-Ab and HBsAg using DBS with a reference test HCV-Ab and HBsAg using serum or plasma and following outcomes were analysed: correlation, regression coefficient, specificity, sensitivity and positive/negative predictive values. We included studies regardless of whether the assay used was commercially available or used an in-house technique, and testing on DBS and plasma/serum did not have to use the same assay. There were no date, geographic or population demographic restrictions, and individuals of all age groups were included.

Screening and data extraction

Two independent reviewers performed title, abstract and full-text review to identify eligible studies. Disagreements were resolved by consent of the reviewers. The references of articles selected for inclusion were also reviewed for additional articles to review. The same data extraction procedure was performed in duplicate for each study and included the following variables: author, publication and study dates, country, percentage of children and adults, age range, gender distribution, type of specimen used for DBS, specimen used as gold standard (plasma or serum), serological assay used, storage conditions and effect of storage conditions. Additional data and clarifications were sought by contacting study authors where necessary.

Risk of bias and quality assessment

The QUADAS-2 tool categories of study design, index and reference test conduct and reporting of patient flow were adapted for use to assess the risk of bias in included studies. In particular, studies where there was consecutive sampling of patients were rated as being at low risk of bias, and a case control design as at high risk of bias. Studies reporting use of a consistent protocol for index and reference testing for each sample and description of patient flow were rated at low risk of bias) while lack of reporting or inconsistent use of a protocol were considered at high risk of bias.

Statistical data analysis

Summary estimates of sensitivity and specificity were generated with a bivariate random effects meta-analysis using maximum likelihood estimate and 95% confidence intervals. We calculated positive (Sensitivity/(1-Specificity) and negative (1-Sensitivity/Specificity) likelihood ratios directly from the pooled sensitivity and specificity. Several studies did not have sufficient quantitative data to contribute to both sensitivity and specificity - for example no samples with a negative reference test. In such cases, we performed a univariate random effects meta-analysis of the sensitivity and/or specificity estimates separately to incorporate studies that did not report estimates for both. We then compared univariate analyses with the result of the bivariate analysis, in order to make complete use of all the available data.

Heterogeneity was assessed visually from forest plots and by reporting an estimate of τ2 corresponding to the variance of the logit-transformed specificity and sensitivity, which can be interpreted as a measure of between-study variability [33]. Stratified analyses were performed by type of assay used for the index test and by storage conditions. Some studies exposed individual samples or subsets of samples to varying storage conditions or used them to define limits of detection. Since these subsets were not necessarily part of the diagnostic accuracy evaluation we included them in the narrative analysis of the impact of storage conditions on the validity of results. Statistical analysis of the data was performed using STATA 14 (StataCorp. 2015. Stata Statistical Software: Release 14. College Station, TX: StataCorp LP).

Results

Included studies

For HBsAg detection, our search identified 521 abstracts, of which 65 full text articles were assessed for eligibility, and 23 [3456] met criteria for inclusion in the review (Fig. 1 & Table 1). Six studies came from Africa [34, 36, 39, 41, 47, 51], nine from Europe [37, 38, 4244, 50, 53, 55, 56], four from the Americas [35, 45, 46, 54], three from Asia [40, 48, 49]and one from Australia [52]. Four studies did not have sufficient data for sensitivity and specificity analysis [40, 43, 44, 53]. Most studies provided limited information on the characteristics of participants. One study only included pregnant women [35], one only children [51] and one only HIV-infected persons [39]. All studies used the same commercial assays for their reference and their index test, respectively. Five studies from the 1980s used Ausria II (Abbott), whereas newer studies used diverse commercial assays, including Enzygnost (Behring) and ARCHITECT (Abbott) (see also Table 1).
Fig. 1
Fig. 1

PRISMA flow chart of studies included in the systematic review of detection of hepatitis B Surface antigen from DBS samples compared to venous blood sampling (plasma/serum)

Table 1

Characteristics of studies included in the systematic review on HBV surface antigen detection from DBS compared to venous blood sampling

Author, Year, Country

Study design

Study pop, Sample size

Storage conditions

DBS collection method

Serum and plasma antigen test

DBS antigen test

Suggested Cut-off

Specificity

Sensitivity

Correlation/Agreement

Effect of storage conditions

Alidjinou 2013 Congo-Brazzaville

Cross-sectional

Attending hepatology clinic, 32

Temperature: Room temperature (25C) Time: 2 months

Whole blood by venipuncture, 30 μl of plasma onto filter paper

Enzygnost ELISA (Siemens)

Enzygnost ELISA (Siemens)

NR

100

100

Spearman correlation coefficient r of 0.89

No significant change

Boa-Sorte

2014 Brazil

Cross-sectional

Pregnant women, 692

Temperature: Refrigerated Time: Less than 5 days

Unclear amount, venipuncture, Schleicher & Schuell 903© filter paper

IMUNOSCREENHBSAG–SS (Mbiolog Diag.), and Murax HB Sag (Murax BioTech Unlmtd.)

IMUNOSCREENHBSAG–SS (Mbiolog Diag.), and Murax HB Sag (Murax BioTech Unlmtd.)

NR

100

100

NR

NR

Brown, 2012, UK

Cross sectional

Unclear

NR

NR

Abbott Architect I2000

Abbott Architect I2000

NR

100 (unclear N)

98% (unclear N)

NR

NR

Farghaly AM 1990 Egypt

Cross sectional

29 HBsAg pos sera 10 sera positive for anticore only 25 sera negative for any marker of HBV

Temperature: Refrigerated (−20 °C)

5 ml by venepuncture, three drops of blood (50-100 μl) on Whatman No.3

Enzygnost HBsAg

Enzygnost HBsAg

NR

100 (25/25)

100 (29/29)

NR

NR

Farzadegan

1978 Iran

Cross-sectional

10 carriers of HBsAg, 10

NR

Capillary (>0.025 ml) Whatman 4

RIA (Austria II, Abbot), Abbot’s Auscell

RIA (Austria II, Abbot), Abbot’s Auscell

NR

NR

100 (10/10)

NR

Samples stored for 1, 3, 7, 14, and 30 days at 4 °C and 37 °C. Best storage temp °4.

Forbi 2010 Nigeria

Cross-sectional

Broad, 300

Temperature: 4C Time: Overnight

Venipuncture, 25 μl of whole blood, Whatman filter paper

Shantest TM- HBsAg ELISA

Shantest TM- HBsAg ELISA

NR

88.6

78.6

NR

NR

Grune 2015 Germany

Cross-sectional

Inpatients 299

Temperature: −20C, 4C or ambient temperature Time: Up to 14 days

Venipuncture, 100 μl whole blood on paper card

Not specified

Not specified

0.15 IU/ml Derived from different sample

99.8

91.7

NR

NR

Halfon 2012

Cohort

Broad, 200

Temperature: -20C (within 48 h of collection) Time: Not specified

Finger prick, 3 drops whole blood on paper card

BV Cobas Taqman (Roche), Abbott Architect

BV Cobas Taqman (Roche), Abbott Architect

2.8 IU/mL

100

98

NR

NR

Kania 2013 Burkina-Faso

Cross-sectional

Attendees of HIV testing center, 218

Temperature: Ambient temperature Time: Not specified

5 spots whole blood (50 μl) on Whatman 903

Determine, SD Bioline, ETI-MAK-4 HBsAg EIA (DiaSorin S.p.A.), Architect HBsAg QT (Abbott)

Determine, SD Bioline, ETI-MAK-4 HBsAg EIA (DiaSorin S.p.A.), Architect HBsAg QT (Abbott)

Optical density: MAPC (mean absorbance of positive control)/2 + 0.3 standard deviations = 0.825.

100

96

Kappa: 0.98

NR

Khan

1996

Pakistan

Cross sectional

Broad, 90

Temperature: 2–8 °C

Time: overnight

Venepuncture, NR

Enzygnost, HBsAg monoclonal II by Behring

Enzygnost, HBsAg monoclonal II by Behring

Cut off value (according to figures: 0.116 mean absorbance value)

100 (86/86)

100 (4/4)

NR

NR

Lukacs Germany 2005

Unclear

70 random samples from newborns (as negative controls), 8 hepatitis b patients

Temperature: NR

Time: NR

Venipuncture, NR

NR

Luminex 100 reader, SREPE

Cut-off median fluorescence intensity 233 (mean + 2 Standard deviations for healthy controls)

NR

100% (8/8)

NR

NR

Lee 2011 Malaysia

Cross-sectional

Patients at a tertiary hospital, 150

Temperature: -20C

Time: NR

Venipuncture, 50 μl whole blood on filter paper

Abbott, Pearson Test

Abbott, Pearson Test

Cut-off point of 1.72

Relative light units

98

97

Pearson value, r = 0.43

NR

Mayer 2012

US

Cross-sectional

 

Temprature: ambient temperatures

Time: Not specified

50 μl on Whatman (8 mm)

ADVIA Centaur 5634 W

ADVIA Centaur 5634 W

 

100

19/19

100

44/44

r = 0.99

Samples stored at ambient temperature and assayed for month four months every week with HBsAg concentration stable

Mendy

2005 The Gambia

Cohort

Broad, 166

Temperature: 30-33C (humid conditions)

Time: Up to 4 weeks

Three to five drops of blood (20 μl) on filter paper

Determine HBsAg (Abbott Laboratories)

Determine HBsAg (Abbott Laboratories)

 

100

96

NR

NR

Mohamed

2013 France

Cohort

Broad, 200

Temperature: Room temperature

Time: 1, 3, 7 and 14 days

Venipuncture

50 μl on 12 mm discs (Whatman)

Chemiluminescent microparticle immunoassay (Abbott)

Chemiluminescent microparticle immunoassay (Abbott)

0.30 +/−0.81 IU/mL.

100

98

r = 0.85

No significant change

Mossner

Denmark

2016

Cohort

404 Prospective patients from hepatitis clinic and blood donors

Temperature: Room temperature

Time 1–5 days

Finger prick, 75 μ on Whatman filter paper

Architect HBs Ab and Architect

HBc Ab assays using the Architect system (Abbott Diagnostics, Delkenheim, Germany)

Architect HBs Ab and Architect

HBc Ab assays using the Architect system (Abbott Diagnostics, Delkenheim, Germany)

The DBS negative patient had serum quantitative level of HbsAg 175 IU/mL.

100% 318/318

98%

84/85

NR

Variation of 24 h to up to 7 d found no difference in stability of samples

Nielsen, 1980, Germany

Cross sectional

110 patients of both sexes older than 6 years attending outpatient department of the General Hospital Barmenda, Cameroon

Temperature

Venipuncture, 50 μl on 16 mm diameter filter paper, Schleicher-Schüll

RIA (Ausria II-test)

RIA (Ausria II-test)

NR

NR

100

11/11

NR

NR

Parkinson, 1996, Alaska

Cross sectional

Alaska Native carriers of HBsAg, volunteer and pregnant females, 10

Temperature and Time NR

Venipuncture, 100 μl applied to 1,5 cm diameter filter paper (No 903, Schleicher &Schuell)

RIA (Ausria II-test)

RIA (Ausria II-test)

Limit of detection approximately 1.25 ng/ml in blood spots

NR

100 (10/10)

NR

Detectable after 8 weeks of storage at ambient temperatures

Ross

2013 Germany

Cross-sectional

Broad, 299

Time and temperature not specified

Venipuncture, 100 μl applied to filter paper (Whatman)

Abbott ARCHITECT HBsAg

Abbott ARCHITECT HBsAg

15 IU/ml (HBsAg)

100

99

r = 0.86, good agreement in Bland-Altmann plot

NR

Villa

Italy

1981

Cross-sectional

Patients attending liver clinic, 24

Temperature: -20C 4C and room temperature.

Time: 1,7, 15, 30, 60, and 180 days

Capillary blood on Whatman filter paper

Ausria II, Ausab, Corab, e-anti e RIA kit (Abbott)

Ausria II, Ausab, Corab, e-anti e RIA kit (Abbott)

Unknown

100

100

NR

Temperature: Storage at room temperature resulted in no significant change compared to samples stored at 4C or -20C.

Time: Storage greater than 15 days negatively affected sensitivity.

Villar

Brazil

2011

Cross sectional

Patients attending HCVlinic, 133

Temperature:-20C, 4–8C, 22–25C, and 22–25C.

Time:1, 7, 14, 21, 42, 63, 112, and 183 days.

either a finger prick or from 70 μl of a whole blood sample onto the 903 Specimen Collection Paper, Whatman Protein Saver Card

ETI-MAK-4 (Diasorin)

ETI-MAK-4 (Diasorin)

Absorbance value 0.115

97

98

NR

Accuracy of DBS samples was stable over 63 days at all temperatures evaluated but after 63 days, accuracy diminished when stored at 22–25C

Zhuang

Australia

1982

Cross-sectional

90 sera selected from serum collection at Fairfield Hospital, Australia

Temperature: incubated at room temperature

Time: 1 night

200 μl on filter-paper (glass-fibre paper, cut into 3x3cm squares)

Ausria II-125

RIA

NR

NR

100

90/90

NR

Storage at 4 °C, ambient temperatures and 37 °C up to 1 month did not lower sensitivity

Zoulek

Sao Tome and Principe

1983

Cross sectional

86 children (48 boys, 38 girls, mean age: 8,3 + − 8,5)

Temperature: stored at 4 °C for 2 weeks, −20 °C for weeks

Time: 2 weeks, then unspecified

Venepuncture 2–4 drops (50-100 μl) on 1 cm filter paper discs

RIA Abbot (unspecified)

RIA Abbot (unspecified)

Lowest concentration analysed 0,1 mg/l

NR

100 24/24

NR

NR

For HCV-Ab detection, our search identified 521 abstracts, of which 101 full text articles were assessed for eligibility, 23 studies met the criteria for inclusion in the review [26, 27, 37, 39, 40, 43, 5571]. Nineteen had sufficient data for inclusion in the quantitative meta-analysis (Fig. 2 & Table 2). Four studies did not have sufficient data for sensitivity and specificity calculations [60, 61, 67] or described a Receiver Operating Characteristic (ROC) curve only [40], and contacting the authors did not yield further information. Two of the 19 studies provided sufficient data to calculate sensitivity but did not have any negative samples to assess specificity [37, 56] (Fig. 2 & Table 2). Of the 23 studies included in the review, most originated from Europe, North America and Australia (16 in total). Four studies were from South America [26, 59, 69, 70] and three from South-East and Central Asia (India [62], Mongolia [60] and Malaysia [40]). All included studies had been published between 1997 to 2017. Most studies used 50 μl to 100 μl of whole blood on filter paper to test for HCV-Ab. Six studies reported using capillary blood samples [26, 55, 57, 58, 63, 66], while others either did not report on this or used venous blood. One study included children [68], however, age ranges or gender for patients were not always reported. Various assays for detection of HCV-Ab in serum and DBS were used (see also Table 2).
Fig. 2
Fig. 2

PRISMA flow chart of studies included in the systematic review on detection of hepatitis C antibody detection from DBS samples compared to venous blood sampling (plasma/serum)

Table 2

Characteristics of studies included in the systematic review on HCV antibody detection from DBS compared to venous blood sampling

Author, Country

Study design

Study pop, Sample size

Storage conditions

DBS collection method

Serum and plasma antibody test

DBS antibody test

Suggested Cut-off

Specificity

Sensitivity

Correlation/Agreement

Effect of storage conditions

Brandao

Brazil 2013

Cross sectional

386 persons, 40 anti-HCV pos, 346 blood donoers HCV non-reactive,

DBS samples air dried at room temperature for 4 h, stored at −20 °C.

Temperature: −20 °C

Time: Not specified

capillary blood by finger prick 75 μl onto Whatman filter paper

MonolisaTM HCV AgAb ULTRA, Bio-Rad (Marnes-la-Coquette, France), and Murex HCV AgAb, Abbott (Kyalami, Republic of South Africa).

MonolisaTM HCV

AgAb ULTRA, Bio-Rad (Marnes-la-Coquette, France), and Murex

HCV AgAb, Abbott (Kyalami, Republic of South Africa).

ROC cutoff: 0.287 nm for Monolisa assay

ROC cut-off for Murex assay 0.238 nm

Derived from the same sample

99.7 (98.4–99.9)

95.9 (93.3–97.8)

97.5 (86.8–99.9)

97.5 (86.8–99.9)

PPV and NPV calculated

Kappa = 0.99 (with ROC cut-off),

stability up to 60 days of storage at room temperature, but less variation at −20 °C

Croom Australia 2006

Cross sectional

103 samples from high risk groups, negative samples from 94 indivdiuals tested at Haematology Lab

Air dried at room temperature, storage at −20 °C, plasma at −20 °C, time of storage 1 week −11 months

Temperature: −20 °C

Time: 1 week −11 months

Venipuncture, 80 μl of each whole blood sample spotted onto Schleicher and Schuell cards (Grade 903)

Monolisa EIA, confirmation test: Murex anti HCV (version 4.0), EIA

Monolisa EIA, confirmation test: Murex anti HCV (version 4.0), EIA

NR

100% (96–100)

108/108

100% (94–100)

75/75

NR

NR

Chevaliez

France 2014

Unclear

529 patients, 183 HCV seronegative, 346 seropositive

NR

NR

EIA HCV assay

EIA HCV assay

0.2

98.9 (96.1–99.7)

99.1 (97.4–99.7)

R = 0.56

NR

Dokubo

US 2014

Cross sectional within a prospective cohort study of those HCV positive being followed

148 participants in a prospective study of HCV

DBS air-dried for 2 h, then sent to another insitute, then stored at −70 °C

Fingerstick on Whatman 903 cards 0.5 ml blood

Standard diagnostics HCV TMA (Norvatis®)

ELISA v3.0(Ortho®).

Standard diagnostics HCV TMA (Norvatis®)

ELISA v3.0(Ortho®).

 

100%

(71/71)

70%

(54/77)

Kappa 0.69

NR

Flores

2016

Brazil

Cross sectional

Participants recruited from ambulatory and general hospital, known HCV/HIV serological status

DBS airdried 4 h, then frozen at -20C

Venipuncture, Whatman filter paper, 3–5 drops (~75 μl)

HCV Murex AB, Diasorin

HCV Murex AB, Diasorin

NR

100%

(99/99)

94%

(213/230)

Spearman correlation r = 0.520

NR

Gruner

2015 Germany

Cross-sectional

Inpatients, 299

Temperature: -20C, 4C or ambient temperature

Time: Up to 14 days

Venipuncture, 100 μl whole blood on paper card

Not specified

Not specified

Derived from different sample

NR

99%

339/343

NR

NR

Kania Burkina-Faso 2013

Cross sectional

218 HIV screening participants, 5 anti-HCV pos, 213 anti-HCV neg

NR

Venipuncture, 5 spots whole blood (50 μl) on Whatman 903

Monolisa HCV-Ab-Ag ULTRA assay (Bio-Rad), further assessment with Inno-Lia HCV Score assay (Innogenetics)

Monolisa HCV-Ab-Ag ULTRA assay (Bio-Rad), further assessment with Inno-Lia HCV Score assay (Innogenetics)

0.439

100% (97,8–100%)

100% (46,3–100%)

kappa: 1,00 (0,93–1,00)

NR

Larrat

France 2012

Cross sectional

One hundred thirteen HCV-positive cases consecutively

recruited 17 HIV/HCV co-infected patients (15%)

DSB dried 24 h at room temperature

Finger prick blood on Whatman card

Monolisa® HCV-Ag-Ab-ULTRA,

Bio-Rad)

Oraquick HCV

CEIA Biorad

0.1

0.2 cEIA

100 (95.8–100) 88/88

100 (95.8–100)

88/88

97.4(92.5–99.1) 110/113

98.2 (93.8–99.5)

111/113

ROC AUC OMT cEIA Biorad: 0.99

ROC AUC FSB cEIA Biorad: 0.918

At 3 days room temperature 3/3 HCV negative samples NR,

ODs lower in HIV co-infected patients

Lee Malaysia 2011

Cross sectional

150 paired samples

Left to dry overnight at room temperature, then stored −20 °C

Venipuncture, 50 μl whole blood on filter paper

Abbott

Abbott

ROC cut off 0.10 RLU

100%

97.3%

ROC curve AUC: 0.99

R = 0.631

NR

Lukacs Germany 2005

Unclear

7 samples from known HCV patients

NR

NR

NR

Luminex

NR

NR

100% 7/7

NR

NR

McCarron

UK 1999

Case control

NR

NR

NR

NR

NR

0.99

1.99

87.5%

100%

100%

97.2%

NR

NR

Marques

Brazil 2012

Cross sectional

21 and 24 HCV reactive patients,

234 individual

and 132 HCV negative

serum stored at −20 °C

Venipuncture, 75 μl whole blood on Whatman paper

Two methods: HCV-Ab Radim, Pomezzia, Italy and ETI-AB-HCVK-4 DiaSorin, Vercelli, Italy

Two methods: HCV-Ab Radim, Pomezzia, Italy and ETI-AB-HCVK-4 DiaSorin, Vercelli, Italy

Radim: manu-facturer’s cut off

ROC curve for DiaSorin EIA using the same sample population

99.5% (98–99.9)

98.9% (96.80–99.55)

97.5%

(86.84–99.94)

88.9% (75.95–96.29)

NR

2–8 degrees C, 20–25 degrees C, and −20 degrees were evaluated, −20 resulted in lowest variation Methods of cut off determination: the receiver operating characteristic curve(AUROC)

Marques

Brazil 2016

Recruited at Viral Hepatitis Lab

99 (59 anti HCV/HCV RNA pos, 40 neg samples)

NR

Venipuncture, 3–5 drops on Whatman filter paper

HCV Ab Radim, Pomezia, Italy

HCV Ab Radim, Pomezia, Italy

 

100%

40/40

94.9%

56/59

  

Mossner Denmark 2016

Cohort

404 Prospective patients from hepatitis clinic and blood donors

Temperature: Room temperature Time 1–5 days

Finger prick, 75 μ on Whatman filter paper

Architect HCV

Ab, using the Architect system (Abbott Diagnostics, Delkenheim, Germany)

Architect HCV

Ab, using the Architect system (Abbott Diagnostics, Delkenheim, Germany)

NR

100%

288/288

97%

112/116

NR

Variation of 24 h to up to 7 d found no difference in stability of samples

Nandagopal India 2014

Unclear

Murex

60 samples

Venipuncture,50 μl of whole blood 903 Whatman card

NR

NR

NR

100 (29/29)

100 (31/31)

Pearson correlation coefficient 0.98

NR

O Brien

US 2001

Multicenter prospective trial (one-arm only)

1286 subjects enrolled in multi-centre study,

Air dry for 30 min, sent in FedEx envelope

Self collected capillary blood with at home kit

NR

HCV Check, Home Access Corp. self use DBS home kit

NR

Several inconclusive and indeterminate results not included in diagnostic accuracy calculations

100%

686/686

99.5% 402/404

NR

NR

Parker

UK 1997

Case control design

80 anti HCV positive samples, 52 negative

569 dB sample fields from South African neonates

Air dry at room temperature before storage at 4 °C

NR, Dried blood field samples

In house IgG ELISA, immunoblot RIBA 3.0

In house IgG ELISA, immunoblot RIBA 3.0

T/N 5.0

T/N10.0

541/569 95.1%

78/80 98%

69/80

86.2%

NR

 

Ross

Germany 2013

Unclear

339 samples

Dried overnight at room temperature

Venipuncture

100 μl of whole blood applied to Whatman 903 filter paper

ARCHITECT

system (Abbott Diagnostics, Delkenheim, Germany).

ARCHITECT

system (Abbott Diagnostics, Delkenheim, Germany).

NR

100% (97.7–100)

160/160

97.8%

(96–100)

175/179

NR

NR

Soulier

France

2017

Cross-sectional

511 patients recruited, with known serostatus for HCV

Temperature:-80

Time: NR

Venipuncture, 50 μl on Whatman filter paper

EIA; aHCV Vitros ECi; Diagnostics, Raritan, New Jersey).

EIA; aHCV Vitros ECi; Ortho-Clinical Diagnostics, Raritan, New Jersey).

NR

312/315

98%

183/186

99%

NR

25 dB samples stored at ambient temperatures (24 °C) for a mean duration (±SD) of 19 ± 1 months

Sensitivity for genotype detection in DBS 84.5%

Sheperd

UK 2013

Cross sectional

19 recently infected

300 chronic carrier

82 resolved infection

DBS stored at 4 °C until use

NR, 50 μl on 903 Whatman Protein Saver cards

ORTHO HCV 3.0 ELISA Test System with Enhanced SAVekit (Ortho Clinical Diagnostics) was used to detect anti-HCV in DBS

NR

Avidity cut-off AI < 30

NR

NR

NR

NR

Tejada-Strop

US 2015

Case control

103 patients with known HCV result, 33 adult patients with chronic HepC with stored samples

-20 °C until 5 years later

NR, 75 μl of whole blood on 12 mm DBS

Two immunoassays, the VITROS anti-HCV IgG chemi-luminescence assay (CIA) and the HCV 3.0 enzyme immunoassay(EIA), both from Ortho Clinical Diagnostics (Rochester, NY),

Two immunoassays, the VITROS anti-HCV IgG chemi-luminescence assay (CIA) and the HCV 3.0 enzyme immunoassay(EIA), both from Ortho Clinical Diagnostics (Rochester, NY),

3.26 CIA

1.5 EIA

Derived in the same sample

Not calculated

CIA 48/52 92%

EIA 90% 47/52

For stored samples CIA: 100% (33/33)

EIA: 32/33 97%

100% (CIA and EIA)

NR

Tuaillon

France 2010

Case control

100 anti HCV pos serum samples and 100 anti HCV neg samples

18 h dried at room temperature, stored at −20°c for 1–8 weeks

NR, 50 μl of whole blood on Whatman 12 mm paper discs

Ortho HCV 3.0 ELISA, immunoblot assay INNO-LIA HCV Score as confirmatory test

Ortho HCV 3.0 ELISA, immunoblot assay INNO-LIA HCV Score as confirmatory test

Threshold value 0.380

98% (97–100)

99% 97–99)

NR

Stability of anti HCV and HCV RNA investigated by varying room temperature exposure 2–12 days until freezing, after 6 days at room temperature ODs > than cut off values

Waterboer

Mongolia 2011

Cross sectional

1022 sexually active women from cross sectional study (response rate 69%)

Room temperature up to 8 h, then −20 °C up to 1 month (serum + DBS)

Venipuncture, Whole blood applied to 5 spots on DBS filter paper cards (Whatman 903)

In house, the

HCV (strain H77, subtype 1a) Core and NS3 proteins

In house, the HCV (strain H77, subtype 1a) Core and NS3 proteins

Sera 1492 (Core)

371 (NS3)

DBS

967 (Core)

310 (NS3)

Not calculable from the data

Not calculable from the data

98% Agreement (kappa 0.94) for Core

96.1% Agreement (kappa 0.90) for NS3

NR

Assessment of study quality and risk of bias (Tables 3 & 4)

Ten studies investigating HBsAg did not use a random or consecutive sampling method [4148, 50, 55, 56]. Only one study reported on blinding of laboratory personnel to results of the reference test while performing diagnostic tests [35]. Overall, the study quality was rated as moderate. For HCV-Ab detection, eight of the included studies used case-control designs [27, 43, 64, 6669, 71] and only two reported consecutive sampling [39, 40]. Only three studies reported blinding of laboratory personnel to results of the reference test [27, 57, 66]. However, most of the other studies (as with HBsAg detection) used and reported a clear and consistent protocol for both reference and index test, so this was not judged as a major cause of bias.
Table 3

Risk of bias in studies included in the systematic review on detection of HB surface antigen

Author

Patient selection

Bias

Index test

Bias

Reference standard

Bias

Flow and timing

Bias

 

Was a case control design avoided?

Consecutive or random sample of patients?

Inappropriate exclusions?

Blinded to reference standard

Could the conduct or interpretation of the index test have introduced bias?

Blinded to index?

Could the reference standard have introduced bias?

There is an appropriate interval between the index test and reference standard?

All patients receive the same reference standard and are included in the analysis?

Alidjinou

NR, but no case control design

UR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Boa-Sorte

No case control design, consecutive recruitment

LR

blinded

LR

blinded

LR

Same reference standard, all patients included in analysis

LR

Brown

Not reported

UR

Not reported

UR

Not reported

UR

Not reported

UR

Farzadegan

Only cases, no consecutive sampling

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Farghaly

Case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Forbi

No case control design

LR

Not blinded, interpretation unbiased

UR

Not blinded, interpretation unbiased

UR

Sampling not reported, same reference standard

UR

Gruner

NR

UR

Not blinded, NR

UR

NR

UR

NR

UR

Halfon

NR, probably case control design

UR

Not blinded, NR

UR

Not blinded, NR

UR

NR

UR

Kania

Consecutive recruitment

LR

Not blinded, interpretation unbiased

UR

Not blinded, interpretation unbiased

UR

Sampling reported

LR

Khan

No case control design

LR

Not reported, unclear whether blinded

UR

Not reported, unclear whether blinded

UR

Sampling not reported

UR

Lee

Consecutive recruitment

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported, same reference standard

LR

Lukacs

Case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling not reported, same reference standard

UR

Mayer

NR, probably case control

UR

Not reported, interpretation unbiased

LR

Not reported, interpretation unbiased

LR

Sampling not reported

UR

Mendy

Case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported

LR

Mohamed

Case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Mossner

Sampling from high-risk and low risk groups

HR

Not blinded, interpretation unbiased

UR

Not blinded, interpretation unbiased

UR

Sampling reported, same reference standard, all patients included in analysis

LR

Nielsen

Only cases

HR

Not reported

UR

Not reported

UR

Sampling not reported

UR

Parkinson

Only cases

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling not reported, same reference standard

UR

Ross

Sampling not reported, probable case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Flow reported

LR

Villa

Case control design

HR

NR

UR

NR

UR

NR

UR

Villar

Case control design

HR

Bias possible, as selective samples by OD values

HR

Not blinded, interpretation unbiased

LR

Sampling reported

LR

Zhuang

No case control design

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling not reported

UR

Zoulek

Unclear, but no case control design, probably random or successive

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling not reported, same reference standard

LR

Abbreviations: LR: low risk, HR: high risk, UR: unknown risk, NR: not reported; shaded: low risk of bias

Table 4

Risk of bias table for HCV antibody

 

Patient selection

Bias

Index test

Bias

Reference standard

Bias

Flow

Bias

 

Was a case control design avoided?

Consecutive or random sample of patients?

Inappropriate exclusions?

 

Blinded to reference standard

Could the conduct or interpretation of the index test have introduced bias?

 

Blinded to index?

Could the reference standard have introduced bias?

 

There is an appropriate interval between the index test and reference standard?

All patients receive the same reference standard?

All patients recruited into the study are included in the analysis?

 

Brandao

No case control design, consecutive sample, no exclusions

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported, same reference standard

LR

Croom

Sampling from high-risk and low risk groups

UR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

All patients included, same reference standard

LR

Chevaliez

NR

UR

NR

UR

NR

UR

NR

UR

Dokubo

No case control, concurrent sampling from a prospective cohort

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported, same reference standard, all patients recruited included in analysis

LR

Flores

Case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported, same reference standard, all patients recruited included in analysis

LR

Gruner

NR

UR

Not blinded, NR

UR

NR

UR

NR

UR

Kania

Consecutive recruitment

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported

LR

Larrat

Consecutive recruitment, but of known cases and known negative controls

HR

blinded

LR

Blinded

LR

Sampling reported, same reference standard

LR

Lee

Consecutive recruitment

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported, same reference standard

LR

Lukacs

NR

UR

NR

UR

NR

UR

Sampling reported, same reference standard

LR

McCarron

Case control, known positive and negative cases from prevalence survey

HR

NR

UR

NR

UR

NR

UR

Marques 2012

No case control design

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling reported, same reference standard

LR

Marques 2016

NR

UR

Not blinded, interpretation unbiased

UR

Not blinded, interpretation unbiased

UR

NR, same reference standard, NR

UR

Mossner

Sampling from high-risk and low risk groups

UR

Not blinded, interpretation unbiased

UR

Not blinded, interpretation unbiased

UR

Sampling reported, same reference standard, all patients included in analysis

LR

Nandagopal

NR

UR

NR

UR

NR

UR

NR

UR

O Brien

No case control design,

LR

Blinded

LR

Blinded

LR

Sampling partly reported, same reference standard

LR

Parker

Case control design

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Sampling partly reported, same reference standard

LR

Ross

Possible case control design, sampling NR

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

Flow reported

LR

Sheperd

No case control design, but partly sampling from patients with known disease

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Soulier

Sampling from high-risk and low risk-groups

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR, same reference standard, NR

LR

Tejada-Strop

Case control

HR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Tuaillon, E

Case control

HR

Blinded

LR

Blinded

LR

Sampling reported, same reference standard

LR

Waterboer, T

No case control

LR

Not blinded, interpretation unbiased

LR

Not blinded, interpretation unbiased

LR

NR

UR

Abbreviations: LR: low risk, HR: high risk, UR: unknown risk, NR: not reported; shaded: low risk of bias

Diagnostic performance

In general, based on the 19 studies evaluated, testing for HBsAg using DBS maintained good accuracy compared to testing using plasma or serum from venipuncture. Reported sensitivity estimates ranged from 79%–100% with a pooled estimate from a meta-analysis of 98% (95%CI 95%–99%), and specificity ranged from 89%–100%, with a pooled estimate of 100% (95%CI 99–100%) (Fig. 3). The positive likelihood ratio was 703 (95%CI 107–4615) and the negative likelihood ratio was 0.02 (95% CI 0.008–0.04). Bivariate and univariate estimates of pooled sensitivity and specificity were similar.
Fig. 3
Fig. 3

Forest plot of sensitivity and specificity of hepatitis B Surface antigen serological diagnosis in DBS compared to venous blood samples

Based on the 19 studies included in the quantitative meta-analysis for HCV antibody, the reported sensitivity for HCV-Ab using DBS ranged from 70% to 100% and specificity ranged from 95 to 100%. The pooled bivariate estimate of sensitivity and specificity was 98% (95%CI 95–99) and 99% (95%CI 98–100), respectively (Fig. 4). The positive likelihood ratio was 361 (95%CI 61–2163) and the negative likelihood ratio 0.02 (95%CI 0.01–0.05). Four included studies reported also measures of agreement with kappa values ranging from 0.87–1 between DBS and venous blood samples [26, 39, 59, 60]. Bivariate and univariate estimates of pooled sensitivity and specificity were similar.
Fig. 4
Fig. 4

Forest plot of sensitivity and specificity of hepatitis C antibody detection in DBS samples compared to venous blood samples

Visual assessment as well τ2 and its p-value showed moderate heterogeneity in the bivariate analysis of studies. To further assess the heterogeneity, we evaluated different parameters with potential to affect accuracy, including assay type and cut-off used, and storage conditions.

Effect of test and cut-off used

There are no standardised cut-offs for HBsAg detection using DBS. Eight of the studies reported a cut-off based on ROC analysis from the same set of samples as the validation set. Several of the included studies noted that the ideal cut-off (as suggested by ROC curves) for determining test positivity should be higher for DBS samples than for plasma or serum samples. Other studies have indicated that this was due to the small sample volume used in DBS (commonly 50 μl). Indeed the one study with low sensitivity (79%) and specificity (89%) [36] only used 25 μl of blood on filter paper (37).

Different assays were used in HCV-Ab detection. Cut-offs varied widely and as no standardized cut-offs exist, investigators for many studies determined their own cut-off via ROC curves using the same set of samples. Nine of the included studies reported on cut-offs used for DBS [26, 27, 40, 59, 60, 6466]. Stratification by type of test or cut-off used was not possible as the number of strata would have been large and the results difficult to interpret. Stratification in a pooled analysis based on amount of blood (50 μl versus >50 μl) or sampling method (venous blood vs capillary blood) showed similar sensitivity and specificity estimates (data not shown).

Effect of storage conditions

For HBsAg detection, the effect of a range of storage conditions was evaluated in six studies, including storage temperature ranging from −20 to 33 °C and storage duration ranging from overnight to 180 days. In general, storage at room temperature or higher (30–33 °C) did not clearly affect accuracy of testing and no decrease in sensitivity was found with prolonged storage at ambient temperatures [34, 4446, 48, 51, 54].

Four studies of diagnostic accuracy for HCV-Ab using DBS samples evaluated different storage conditions in a subset of samples that did not contribute to the diagnostic accuracy evaluations. In one study, three out of three previously negative samples exceeded cut-off values (i.e. would have been interpreted as positive) after storage for 3 days at room temperature [66]. Similarly, one of the included studies showed that after 6 days of storage at room temperature, the cut-off values were exceeded and previously negative samples became positive [63]. Two studies showed relative stability at room temperature for up to 60 days, but found that storage at −20 °C was associated with less variation in quantitative values [26, 59].

One of the 19 studies contributing to the diagnostic accuracy calculations kept study samples at room temperature for more than 24 h [55]. A pooled analysis stratifying studies according to whether samples had been left at room temperature for longer than 4 h or not did not find any difference in performance and so did not explain the heterogeneity observed in the meta-analysis (data not shown).

Sensitivity analysis

In a sensitivity analysis, we found no difference in estimates of diagnostic performance between those studies that reported consecutive sampling and those with case-control study design (data not shown).

Discussion

To our knowledge, this is the first comprehensive and systematic review to summarize the utility of DBS for HCV-Ab and HBsAg testing. Overall, we found very good diagnostic accuracy and precision for detection of HCV-Ab and HBsAg using DBS samples. These findings were based on a larger number of studies compared to the companion systematic reviews undertaken for HBV DNA and HCV RNA using DBS [30] and were rated as moderate quality. This provides support for the use of DBS from capillary blood for diagnostic testing of HBV or HCV, especially in community settings and hard to reach populations where there is limited access to venipuncture or inadequate laboratory infrastructure to prepare or transport plasma samples or limited access to rapid diagnostic tests.

The WHO 2017 testing guidelines recommended the use of a single quality-assured serological assay (i.e either a laboratory-based EIA or RDT to detect HBsAg and anti-HCV that meet minimum performance standards, and where possible delivered at the point of care to improve access and linkage to care and treatment. The guidelines also made a conditional recommendation to consider the option of DBS specimens for HBsAg and HCV-Ab serology testings in settings where there are no facilities or expertise to take venous whole blood specimens; or RDTs are not available or their use is not feasible, or there are persons with poor venous access (e.g. drug treatment programmes, prisons) [29]. This recommendation was rated as conditional mainly because of the slightly lower accuracy using DBS compared to venous blood sampling and uncertainty regarding the generalizability of studies included in this review (in particular regarding impact of different storage and transport conditions).

There were several limitations to this review. First, we did not include studies in languages other than English and no unpublished data from laboratories were included. Second, while only a few studies were rated as having a low risk of bias, overall the quality of evidence from studies was rated as moderate. Third, most studies did not use fresh samples and no uniform protocol was applied. Fourth, there is also a risk of overestimation of pooled sensitivity and specificity by including studies that applied cut-off levels derived from the same study population. No stratified analysis was possible for the type of test within this review. Therefore, we are unable to recommend the use of certain commercial tests over others using DBS testing of HCV-Ab or HBsAg or to suggest a cut-off specific to a test that should be used for DBS samples.

Several studies found that when assessing stability in separate sample sets samples became false-positive after longer exposure at ambient temperatures for both HCV-Ab [27, 66] and HBsAg [44, 45]. Further data is needed to understand the stability of DBS with different environmental conditions (i.e. temperature and humidity). The review highlights the need for standardized validation of specific tests with DBS.

While some studies published detailed protocols on how to collect and analyze DBS [37], no manufacturers to date have provided instructions on how to use their assays with DBS (including processing methods and possibly different cut-offs). There is a need for manufacturers to validate their assays and provide instructions for the use of DBS even if no claim for regulatory approval is made.

Implementation of DBS and future work:

Consideration of the use DBS sampling for HBV and C serological or nucleic acid testing or both will depend on the health-care setting and infrastructure, and epidemiological context. If good-quality RDTs are available that can be performed using capillary blood then the focus may be more on prioritizing DBS for NAT testing of HBV DNA and HCV RNA. However, if RDTs are not available and there are no facilities or expertise to take venous blood samples, then DBS testing may be equally important to increase access to serological testing as well as NAT. Conveniently, both could be performed from the same specimen. A further situation where DBS may be applicable is where large numbers of individuals are being tested simultaneously. DBS may also be useful where polyvalent screening for multiple diseases is done, but where multiplex RDTs for this purpose are not available or are more costly. The adoption of DBS sampling in a hepatitis testing programme requires the availability of a centralized laboratory competent at handling and processing this sample type. The current lack of validation of assays on this sample type for HBsAg or HCV-Ab and manufacturer’s guidance on the optimal pre-analytical treatment of specimens makes quality control challenging.

Further data is also needed on demonstrating the stability of DBS with individual tests under different field transport and storage conditions likely to be encountered in low resource settings. as well as on the type of test used and how long DBS samples can be left at different temperature or humidity levels. Since DBS requires only a small sample of blood to maintain sensitivity, a higher analytical cut-off may be required to maintain sensitivity and overcome variability at the lower end of the dynamic range of the test as compared to higher volume plasma samples – even if in our limited stratified analysis on this we did not find an effect of blood volume on diagnostic accuracy of HBV or HCV. Further insight can also be gained from individual patient analyses of the existing data, which was beyond the scope of this review.

Conclusion

While diagnostic accuracy of DBS for HCV-Ab and HBsAg testing is adequate in studies included in this review, lack of standardization of testing protocols and uncertainty about their use in field conditions and the appropriate assay cut-offs limits the wider application of DBS.

Declarations

Funding

Funding for this study was provided by the World Health Organization, Global Hepatitis Program. Publication of this article was funded by the World Health Organization.

Availability of data and materials

Not applicable.

About this supplement

This article has been published as part of BMC Infectious Diseases Volume 17 Supplement 1, 2017: Testing for chronic hepatitis B and C – a global perspective. The full contents of the supplement are available online at https://bmcinfectdis.biomedcentral.com/articles/supplements/volume-17-supplement-1.

Authors’ contributions

BL, JCo and CMD designed the study according to PICO questions stated during the WHO guideline process. BL, JCa, CP designed the search strategy and performed the search, and data extraction for HCV-Ab DBS studies. JCo, JCh, NG, AN and JG performed the search, screening and data extraction for the HBV surface antigen DBS studies. TR, AI, ET PvdP and PE provided important design and technical insight. BL and JCo performed the meta-analyses. BL, CMD and PE wrote the manuscript. All authors helped with revisions and the final version of the manuscript.

Ethics approval and consent to participate

Not applicable

Consent for publication

Not applicable

Competing interests

The authors declare that they have no competing interests.

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Authors’ Affiliations

(1)
Division of Infectious Diseases, Department of Medicine II, Medical Center – University of Freiburg, Faculty of Medicine, University of Freiburg, Germany, Freiburg, Germany
(2)
Centre for Chronic Immunodeficiency, Medical Center – University of Freiburg, Faculty of Medicine, University of Freiburg, Germany, Freiburg, Germany
(3)
Department of Infectious Diseases, University of Pennsylvania, PA, Philadelphia, USA
(4)
FIND, Geneva, Switzerland
(5)
Medicines sans Frontières, Access Campaign, Paris, France
(6)
School of Medicine, Boston University, Boston, USA
(7)
Global Hepatitis Programme, HIV Department, World Health Organization, Geneva, Switzerland
(8)
Sargent College, Boston University, MA, Boston, USA
(9)
Pathogenesis and Control of Chronic Infections UMR 1058 INSERM/Université Montpellier/Etablissement Français du Sang, INSERM, 34394 Montpellier Cedex 5, France
(10)
Centre Hospitalier Universitaire (CHU) de Montpellier, Département de bactériologie-virologie, Montpellier, France
(11)
Department of Medicine II, Medical Center – University of Freiburg, Faculty of Medicine, University of Freiburg, Freiburg, Germany

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