Evaluation of two commercial human T-cell lymphotropic virus western blot (immunoblot) kits with problem specimens

Abstract

We evaluated two commercial human T-cell lymphotropic virus (HTLV) Western blot (WB; immunoblot) kits, Cambridge Biotech Corp. (CBC) and Diagnostic Biotechnology Ltd. (DBL). Both methods employ HTLV type I (HTLV-I) viral lysate and rgp2l. The DBL WB kit also distinguishes between HTLV-I and HTLV-II antibodies, using an HTLV-I-specific and an HTLV-II-specific recombinant. Fifty weakly reactive HTLV-IIpositive plasma specimens which were falsely negative with the Abbott enzyme immunoassay (EIA) and 50 Ortho EIA false-positive samples were selected to determine sensitivity and specificity. The sensitivities of the CBC and the DBL WB kits were 90 and 68%, respectively. All positive samples reacted with rgp2l in both kits, but some did not display core bands. Five samples were typed as HTLV-I and four were typed as dual infection by the DBL WB kit. The specificities of the CBC and DBL kits were 48 and 70%o, respectively. The most prevalent WB reaction with the negative samples was with the core protein, p19, followed by p24 and p28 for CBC and rgp2l and p28 for DBL. DBL had two false-positive interpretations, and CBC had none. rgp2l was the most sensitive antigen in both kits for the weakly reactive HTLV-II samples. If all samples not reacting with this protein were interpreted as VB negative, regardless of other bands, the specificity would improve to 90% for CBC and 86% for DBL.

Enhanced specificity of truncated transmembrane protein for serologic confirmation of human T-cell lymphotropic virus type 1 (HTLV-1) and HTLV-2 infections by western blot (immunoblot) assay containing recombinant envelope glycoproteins

“…Our group has further demonstrated that the lack of reactivity to rgp46 2 in specimens from Seminole Indians was not due to sequence variation but, rather, reflected lower titers of HTLV-2 antibodies (15). A larger study with HTLV enzyme immunoassay-negative specimens (PCR positive for HTLV-2) has further revealed that specimens containing low titers of antibodies to HTLV-1/2 also can be missed by this test (5). We therefore strongly recommend that all diagnostic specimens with reactivity to p24 and GD21-I continue to be referred to as HTLV-positive untypeable, requiring further testing by more sensitive assays.…”

Laboratory performance in HTLV‐I/II analysis

“…Before each MPEP shipment, all donor samples were tested independently by two laboratories to confirm retroviral serologic status by using all available HTLV and HIV EIA test kits licensed by the FDA and all available WB kits. In addition, each HTLV‐I/II antibody‐positive donor sample was tested by RIPA and, since 1994, by type‐specific WB and indirect immunofluorescent (IIF) antibody assays capable of distinguishing HTLV‐I from HTLV‐II 3,4 . Preshipment HTLV‐positive serologic status of donor samples was based on repeatedly reactive results on all FDA‐licensed HTLV EIAs and demonstration of antibody to both core and envelope antigens in HTLV WBs and/or RIPA 1 .…”

Diversidade molecular do envelope e carga proviral do vírus linfotrópico de células T humanas tipo 2 (HTLV-2) em amostras indeterminadas pelo teste Western-Blot

“…This would have increased the specificity of the HTLV WB 2.4 kit to 86%. They concluded that the indeterminated WB are caused by lack of sensitivity and specificity of the WB antigen and poor interpretation criteria, not by variances of the specimens [Gallo et al 1994].…”

“…É possível considerar que o sistema imune é um fator importante e podem resultar as amostras de soro em indeterminado, possivelmente pela menor produção de anticorpos anti-HTLV-2(Bassini et al, 2006). No entanto, nossos resultados indicam que a contagem células T não influenciou na presença dos resultados indeterminados, mesmo em indivíduos infectados pelo HIV-1 Gallo et al (1994). constataram que o teste de WB tinha uma sensibilidade de 68%, com fracas bandas nos resultados de HTLV-2 positivos e uma especificidade de 70% com os negativos.…”

“…constataram que o teste de WB tinha uma sensibilidade de 68%, com fracas bandas nos resultados de HTLV-2 positivos e uma especificidade de 70% com os negativos. Apesar de que todos os positivos reagiram com rgp21, a presença do antígeno p24 no kit não foi suficientemente para reagir com todos os aspectos positivos e 11 amostras de HTLV-2 reagiram com a proteína do envelope(Gallo et AL. 1994).…”