The INNO-LIA® HTLV I/II Score (Fujirebio, Japan) was used as a method for confirmation and differentiation of viral types, following the manufacturer’s protocol.
Inno lia htlv 1 2 score
Manufactured by Fujirebio
Sourced in Japan
The INNO-LIA® HTLV I/II Score is a laboratory equipment product developed by Fujirebio. It is designed to detect and differentiate antibodies to human T-cell lymphotropic virus type I (HTLV-I) and type II (HTLV-II) in human serum or plasma samples.
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11 protocols using inno lia htlv 1 2 score
1
HTLV I/II Viral Typing Protocol
2
HTLV-1/2 Antibody Screening and Confirmation Protocol
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Samples were initially screened for HTLV1/2 antibodies using one of the following three commercially available ELISA assays, including Wantai HTLV-1/2 antibody ELISA kits (Wantai BioPharm, China), Murex HTLV I+II (Diasorin S.p.A., United Kingdom), and Foresight HTLV-1/2 antibody ELISA kits (Acon Biotech, China). Reactive specimens underwent retests by the same assay, and any one of the two-round retests that were reactive would be defined as an initially anti-HTLV-1/2 reactive specimen and was sent to the NCCL for further confirmation.
Two different serologic assays, namely, Elecsys HTLV-I/II (Roche Diagnostics, Germany) and Murex HTLV I+II (Diasorin S.p.A., UK), were performed on one sample simultaneously in NCCL. Any sample that reacted to one of the two assays was confirmed and discriminated by the line immunoassay (INNO-LIA HTLV I/II Score, Fujirebio, Japan).
Two different serologic assays, namely, Elecsys HTLV-I/II (Roche Diagnostics, Germany) and Murex HTLV I+II (Diasorin S.p.A., UK), were performed on one sample simultaneously in NCCL. Any sample that reacted to one of the two assays was confirmed and discriminated by the line immunoassay (INNO-LIA HTLV I/II Score, Fujirebio, Japan).
3
Nationwide HTLV-1 Infection Screening
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All the specimens were collected from eligible donors from blood banks/centers in 28 provinces and Shenzhen city in China from January 2019 to December 2021, and 12,083 samples from the Shandong blood hematopoietic stem Cell Bank were also included. The initially reactive samples were delivered at 2–8°C or frozen conditions to the National Center for Clinical Laboratories (NCCL) for confirmation. According to the confirmatory process (Figure 1 ), all the initially reactive samples were simultaneously tested by two screening assays in the NCCL. Samples reactive to any one of the two tests were further tested by a line immunoassay (LIA) (INNO-LIA HTLV I/II score, Fujirebio, Japan).
Samples with limited plasma volume that were not sufficient for all assays' tests were excluded from our study. Enrolled samples with whole blood available were further quantified by qPCR established in our laboratory. Samples that can be detected using qPCR were also amplified for the LTR region of HTLV-1 by nested PCR. In addition, blood donors with positive results were recommended to collect blood samples from their relatives and test them using the same detection process. The positive donors were also asked to fill out a questionnaire about their personal experiences of probably being infected with HTLV-1/2 and some of the HTLV-1-related syndromes they have suffered.
Samples with limited plasma volume that were not sufficient for all assays' tests were excluded from our study. Enrolled samples with whole blood available were further quantified by qPCR established in our laboratory. Samples that can be detected using qPCR were also amplified for the LTR region of HTLV-1 by nested PCR. In addition, blood donors with positive results were recommended to collect blood samples from their relatives and test them using the same detection process. The positive donors were also asked to fill out a questionnaire about their personal experiences of probably being infected with HTLV-1/2 and some of the HTLV-1-related syndromes they have suffered.
4
HTLV-1/2 Antibody Screening and Confirmation Protocol
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Samples were initially screened for HTLV1/2 antibodies using one of the following three commercially available ELISA assays, including Wantai HTLV-1/2 antibody ELISA kits (Wantai BioPharm, China), Murex HTLV I+II (Diasorin S.p.A., United Kingdom), and Foresight HTLV-1/2 antibody ELISA kits (Acon Biotech, China). Reactive specimens underwent retests by the same assay, and any one of the two-round retests that were reactive would be defined as an initially anti-HTLV-1/2 reactive specimen and was sent to the NCCL for further confirmation.
Two different serologic assays, namely, Elecsys HTLV-I/II (Roche Diagnostics, Germany) and Murex HTLV I+II (Diasorin S.p.A., UK), were performed on one sample simultaneously in NCCL. Any sample that reacted to one of the two assays was confirmed and discriminated by the line immunoassay (INNO-LIA HTLV I/II Score, Fujirebio, Japan).
Two different serologic assays, namely, Elecsys HTLV-I/II (Roche Diagnostics, Germany) and Murex HTLV I+II (Diasorin S.p.A., UK), were performed on one sample simultaneously in NCCL. Any sample that reacted to one of the two assays was confirmed and discriminated by the line immunoassay (INNO-LIA HTLV I/II Score, Fujirebio, Japan).
5
Nationwide HTLV-1 Infection Screening
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All the specimens were collected from eligible donors from blood banks/centers in 28 provinces and Shenzhen city in China from January 2019 to December 2021, and 12,083 samples from the Shandong blood hematopoietic stem Cell Bank were also included. The initially reactive samples were delivered at 2–8°C or frozen conditions to the National Center for Clinical Laboratories (NCCL) for confirmation. According to the confirmatory process (Figure 1 ), all the initially reactive samples were simultaneously tested by two screening assays in the NCCL. Samples reactive to any one of the two tests were further tested by a line immunoassay (LIA) (INNO-LIA HTLV I/II score, Fujirebio, Japan).
Samples with limited plasma volume that were not sufficient for all assays' tests were excluded from our study. Enrolled samples with whole blood available were further quantified by qPCR established in our laboratory. Samples that can be detected using qPCR were also amplified for the LTR region of HTLV-1 by nested PCR. In addition, blood donors with positive results were recommended to collect blood samples from their relatives and test them using the same detection process. The positive donors were also asked to fill out a questionnaire about their personal experiences of probably being infected with HTLV-1/2 and some of the HTLV-1-related syndromes they have suffered.
Samples with limited plasma volume that were not sufficient for all assays' tests were excluded from our study. Enrolled samples with whole blood available were further quantified by qPCR established in our laboratory. Samples that can be detected using qPCR were also amplified for the LTR region of HTLV-1 by nested PCR. In addition, blood donors with positive results were recommended to collect blood samples from their relatives and test them using the same detection process. The positive donors were also asked to fill out a questionnaire about their personal experiences of probably being infected with HTLV-1/2 and some of the HTLV-1-related syndromes they have suffered.
6
HTLV-I/II Screening and Confirmation Protocol
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All specimens were collected from eligible donors from blood banks/centers in 26 provinces in China between January 2016 and April 2019. The RR samples were delivered at 2–8°C or frozen to the NCCL for confirmation. The RR samples were also HBsAg, anti-HCV, anti-HIV, and anti-TP negative by both two screening ELISAs in the enrolled blood centers. According to the confirmatory process (Figure 1 ), all RR samples were simultaneously subjected to four screening assays in the NCCL. Samples reactive to any one of the four assays were further confirmed by a LIA (INNO-LIA HTLV I/II score, Fujirebio, Japan) and WB (MP HTLV Blot 2.4, MP Biomedicals, Singapore).
Samples with limited plasma volumes that were not enough for all assays were excluded from our study. Enrolled samples with a positive result either by WB or LIA were classified as positive. Positive or indeterminate specimens with whole blood available were further quantified by qPCR in our lab. In addition, blood donations with an indeterminate result were recollected from the same donor, and retested by the same detection process.
Samples with limited plasma volumes that were not enough for all assays were excluded from our study. Enrolled samples with a positive result either by WB or LIA were classified as positive. Positive or indeterminate specimens with whole blood available were further quantified by qPCR in our lab. In addition, blood donations with an indeterminate result were recollected from the same donor, and retested by the same detection process.
7
HTLV Infection Verification Protocol
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In the follow-up year, the coordinating center of HEMOSUL screened all serum samples using chemiluminescent microparticle immunoassay (CMIA) (Abbott Architect rHTLV I/II kit and ARCHITECT i2000SR equipment) for the presence of anti-HTLV-1/2 antibodies. It was considered positive when samples had a cut-off ≥ 1, following the manufacturer’s instructions. The grey zone considering a cut-off ranging from 0.9 to 1.0, is configured in the equipment by HEMOSUL. Positive samples were confirmed by HTLV-1/2 INNO-LIA assay (Fujirebio INNO-LIA HTLV I/II Score).
Our group retested all the positive serum samples, provided by HEMOSUL, using an enzyme-linked immunosorbent assay (ELISA) commercial kit for the presence of anti-HTLV-1/2 antibodies (Murex HTLV I + II—DiaSorin), following the manufacturer’s instructions. Positive samples were repeatedly tested and confirmed by HTLV-1/2 Western Blot (WB) assay (MP Diagnostics HTLV BLOT 2.4—Singapore). HTLV infection was defined as a repeatedly positive ELISA and positive WB (Fig.1 ).
Our group retested all the positive serum samples, provided by HEMOSUL, using an enzyme-linked immunosorbent assay (ELISA) commercial kit for the presence of anti-HTLV-1/2 antibodies (Murex HTLV I + II—DiaSorin), following the manufacturer’s instructions. Positive samples were repeatedly tested and confirmed by HTLV-1/2 Western Blot (WB) assay (MP Diagnostics HTLV BLOT 2.4—Singapore). HTLV infection was defined as a repeatedly positive ELISA and positive WB (Fig.
8
HTLV Diagnosis and Viral Load Quantification
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Blood samples were collected in advance for all HTLV studies to avoid recitation of the patients and following the diagnostic algorithm described (Figure 2 ). Antibodies for HTLV-1 were screened by quimioluminiscence [rHTLV I/II (CMIA, Architect®, Abbott)] until 2018 and since 2019 by electroquimioluminiscence Elecsys® HTLV I/II (Roche, Mannheim) and reactive samples were tested by LIA (INNO-LIA® HTLV I/II Score, Fujirebio, Japan) as a confirmatory test following the manufacturer's instructions. A qualitative PCR was performed for untyped virus results. Confirmed HTLV-1 samples were further tested for pVL for HTLV-1 in peripheral mononuclear cells (PBMCs) by an in-house real-time quantitative PCR (qPCR) as previously described (33 (link)). The assay had a detection limit of 400 HTLV-1 copies/106 PBMCs, with a broad range of quantitation (2.6 log10 to −6 log10) and a total error accepted of 0.5 log10. Both molecular studies were performed at the Virology Unit in the Hospital Garrahan.
9
HTLV Serological Screening Protocol
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Samples from which it was not possible to obtain leukocytes for DNA extraction and those negative in qPCR, in order to rule out possible cases of false negatives, were submitted to the complementary tests using in-line immunoassay INNO-LIA HTLV-I/II Score (Fujirebio, Tokyo, Japan) and enzyme immunoassay HTLV Blot 2.4 kit (MP Diagnostics, Singapore, Republic of Singapore).
10
Serological and Molecular Detection of HTLV-1/2
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Immunoenzymatic assays (Murex HTLV-I+II, DiaSorin, Dartford, UK) were used to detect anti-HTLV-1/2 antibodies in the tested samples following the manufacturer's instructions. Samples with positive results were subjected to line immunoassays (INNO-LIA® HTLV I/II Score, Fujirebio, Japan) and real-time PCR (Applied Biosystems Step One Plus Real Time PCR) to confirm infection and differentiate the viral type, respectively.
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