Simultaneous phenotyping of five Rh red blood cell antigens on a paper-based analytical device combined with deep learning for rapid and accurate interpretation.

Both the ABO and Rhesus (Rh) blood groups play crucial roles in blood transfusion medicine. Herein, we report a simple and low-cost paper-based analytical device (PAD) for phenotyping red blood cell (RBC) antigens. Using this Rh typing format, 5 Rh antigens on RBCs can be simultaneously detected and macroscopically visualized within 12 min. The proposed Rh phenotyping relies on the presence or absence of hemagglutination in the sample zones after immobilizing the antibodies targeting each Rh antigen. The PAD was optimized in terms of filter paper type, antibodies, and distance of the visualization zone. In this study, the optimal conditions were Whatman filter paper Grade 4; anti-D, -C, -E, -c, and -e antibodies; RBC suspension of 30%; and a visualization zone of 1 cm above the sample zone. The accuracy of simultaneously phenotyping the five Rh RBC antigens in the blood samples (n = 4692) was 99.19%, comparable with the accuracy of the gold-standard tube method used by blood bank laboratories in several regions of Thailand. Furthermore, decision making based on this method can be assisted by deep learning. After implementing a two-stage objective detection algorithm (YOLO v4-tiny) and classification model (DenseNet-201), the ambiguous images (n = 48) were interpreted with 100% accuracy. The PAD integrated with customized-region convolutional neural networks can reduce the interpretation discrepancies in RBC antigen phenotyping in any laboratory.

Human T-cell lymphotropic virus type I and II seroprevalence among volunteer blood donors in Thailand.

Human T-cell lymphotrophic virus type I and II (HTLV-I/II) are closely related but distinct retroviruses that can infect humans. Both the viruses can be transmitted via transfusion of contaminated blood components. HTLV pre-transfusion screening is not mandatory in Thailand until now. Current epidemiological data for HTLV prevalence is still lacking since the past surveys were done more than a decade ago. The main objective of this study was to determine the seroprevalence of HTLV-I/II among voluntary blood donors in Thailand. 11,057 volunteer blood donors were screened for HTLV-I/II antibodies using the ARCHITECT rHTLV-I/II chemiluminescent immunoassay (CLIA). Initial-reactive (IR)  samples were subjected to repeat duplicate testing and were also sent for confirmatory testing at Korean Red Cross Society (KRC), Seoul or National Serology Reference Laboratories (NRL), Australia using alternate HTLV serological assays and immunoblot and/or specific nucleic acid testing respectively. Out of 11,057 plasma samples, 10,080 were low-risk seronegative donors and 977 were first-time/high-risk donors. Twenty of 24 IR samples were repeatedly reactive (RR) in low-risk seronegative donors group. On confirmatory testing of these 24 IR by immunoblot, 13 indeterminate and 11 negative results were observed. One out of 977 samples from first-time/high-risk donors was RR for anti-HTLV-I/II antibodies. This sample was co-reactive for HBsAg, but negative for HTLV by EIA or in-house HTLV-I qPCR. The ARCHITECT rHTLV-I/II assay exhibited a specificity of 99.93% in low-risk donors and 99.90% among high-risk donors. This study concluded that HTLV-I/II prevalence is low among blood donors in Thailand. But periodic surveillance should be continually conducted to ensure high blood safety standards in the country.

A simple and low-cost portable paper-based ABO blood typing device for point-of-care testing.

ABO blood group is the most important blood type system for transfusion medicine. A paper-based analytical device (PAD) for ABO blood typing has been proposed. The device was composed of Whatman No. 113 paper, an absorbent gel pad, and a 3D-printing cassette. The 3D-printing cassette contained two circular holes for display of letters "A" and "B" on the PAD. Whole blood was dropped onto hydrophilic letters A and B on the PAD, in which the anti-A and anti-B were pre-immobilized, respectively. An absorbent gel pad was used to adsorb excess blood sample and washing solution during the washing step. The particle size of agglutinated red blood cells (RBCs) could not be eluted out of the paper by the elution solution. In contrast, non-agglutinated RBCs were washed out by means of elution solution. The devices could be used for real blood samples in a wide range of hematocrit levels, 21-59%. Unknown blood group samples (n = 500) were identified by the developed device and the results were compared with the conventional method, revealing 100% accuracy. Because of its compact size with low-cost fabrication, the portable ABO blood typing device has great potential for point-of-care testing, particularly in developing countries.

Performance evaluation of the Elecsys syphilis assay for the detection of total antibodies to Treponema pallidum.

Syphilis is a health problem of increasing incidence in recent years that may have severe complications if not diagnosed and treated at an early stage. There are many diagnostic tests available for syphilis, but there is no gold standard, and diagnosis therefore usually relies upon a combination of tests. In this multicenter study, we evaluated the treponemal Elecsys syphilis assay for use in the diagnosis of syphilis in routine samples, i.e., when syphilis is suspected or during antenatal or blood donation screening. The sensitivity and specificity of the Elecsys syphilis assay were compared head to head with those of other treponemal assays used in routine clinical practice and were assessed in potentially cross-reactive samples from patients with Epstein-Barr virus, HIV, and Lyme disease. In a total of 8,063 syphilis-negative samples collected from routine diagnostic requests and blood donations, the Elecsys syphilis assay had a specificity of 99.88%. In 928 samples previously identified as syphilis positive, the sensitivity was 99.57 to 100% (the result is presented as a range depending on whether four initially indeterminate samples are included in the assessment). The specificity of the Elecsys syphilis assay in patients with other infections was 100%; no false-positive samples were identified.

Occult hepatitis B infection in blood donors from South East Asia: molecular characterisation and potential mechanisms of occurrence.

OBJECTIVE: To investigate the molecular basis of occult hepatitis B virus (HBV) infection (OBI) in Asian blood donors. DESIGN: OBI donors from Hong Kong, Malaysia, Singapore, Taiwan and Thailand were tested for HBV serological markers, and strains were molecularly characterised. RESULTS: Among 138 confirmed OBI carriers (median age 47 years), HBV genotypes B and C were dominant (60% and 34%, respectively) in agreement with the genotype distribution in chronically infected donors in the region. Viral load ranged between unquantifiable and 3670 IU/ml (median 11 IU/ml). Eleven per cent of OBIs showed an unusual anti-HBs-only serological profile without evidence of past vaccination for most of these individuals. Occult HBV strains showed a higher genetic diversity than strains from matched hepatitis B surface antigen (HBsAg)+ donors, irrespective of genotype. No unique genetic signature or evidence of reduced replication competence was found. Mutations in the vicinity of the pre-S2/S splice donor site were common in OBI(B) (44%) and OBI(C) (36%) strains. S regions from four OBI cases were transfected in HuH7 cells. Results showed limited HBsAg secretion and suggested that mutations disrupting the splice donor site structure may affect pre-S2/S mRNA splicing. CONCLUSIONS: There is indirect evidence that incomplete immune control is involved in the occurrence of OBI in Asian blood donors infected with genotypes B and C as observed in Europe with genotype A2 but to a lower extent than with genotype D. A post-transcriptional mechanism may play a role in HBsAg expression in some OBIs irrespective of HBV genotype.

One-year experience of nucleic acid technology testing for human immunodeficiency virus Type 1, hepatitis C virus, and hepatitis B virus in Thai blood donations.

BACKGROUND: Blood donations collected at the National Blood Center, the Thai Red Cross Society, Bangkok, in 2007 were tested by nucleic acid amplification technology (NAT) using the Chiron TIGRIS/Procleix Ultrio test and the Roche cobas s 201/cobas TaqScreen multiplex (MPX) test. STUDY DESIGN AND METHODS: The sensitivity, specificity, and robustness were determined by testing 486,676 seronegative blood donations. Samples from each day of collection were divided into two sets; the odd-numbered samples were tested individually on the TIGRIS and the even-numbered samples were tested in pools of 6 on the cobas s 201. The status of reactive samples was confirmed by duplicate testing of samples from the plasma bag to calculate the test specificity. Reactive samples were tested on the alternate system and followed up. RESULTS: The analytical sensitivity of both systems met the 95% limits of detection claimed by the respective package inserts. No cross contamination was seen with either system. Test specificity was 99.93 and 99.90% for the Procleix Ultrio and cobas TaqScreen tests, respectively. The NAT yield rates for human immunodeficiency virus Type 1 (HIV-1), hepatitis C virus (HCV), and hepatitis B virus (HBV) were 1:97,000, 1:490,000, and 1:2800, respectively. Several occult HBV donors, the majority of whom were detected by both tests, were also identified. The HIV-1 and HCV window cases were detected with both tests. CONCLUSION: The performances of the systems and tests indicated that both were acceptable for routine NAT by the National Blood Center, the Thai Red Cross Society. However, the Procleix Ultrio test appeared to be less sensitive than the cobas TaqScreen test for HBV.