Anti-BSA antibodies are a major cause of non-specific binding in insulin autoantibody radiobinding assays

Williams, A. J. K.; Curnock, Rachel; Reed, Charles R.; Easton, Peter; Rokni, Saba; Bingley, Polly J.

doi:10.1016/j.jim.2010.09.009

Cited by 13 publications

(12 citation statements)

References 10 publications

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“…The assay was, however, unable to distinguish between the IA + and IA - donors with many samples producing high ECL signal independent of antibody status (Figure 7). This result was not entirely unexpected as others have reported non-specific interfering factors when assessing IAA in human samples [29]. As ECL-based assays have previously been successfully used in human plasma and serum samples for other analytes, we were confident that the ECL platform could still allow for the sensitive detection of human antibodies against insulin.…”

Section: Resultssupporting

confidence: 72%

“…Clarification of human samples with SA-beads to remove non-specifically bound materials significantly decreased background while maintaining specific signal. This clearance step overcomes challenges present in the RBA where assay components including BSA [29] reduce the specificity of that IA assay format. The ECL assay requires the same sample volume per well used in RBA but does not require the generation of a standard curve using known positive samples (Table 3) [30].…”

Section: Discussionmentioning

confidence: 99%

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Antibodies against insulin measured by electrochemiluminescence predicts insulitis severity and disease onset in non-obese diabetic mice and can distinguish human type 1 diabetes status

Swafford

Shafer‐Weaver

et al. 2011

J Transl Med

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BackgroundThe detection of insulin autoantibodies (IAA) aids in the prediction of autoimmune diabetes development. However, the long-standing, gold standard 125I-insulin radiobinding assay (RBA) has low reproducibility between laboratories, long sample processing times and requires the use of newly synthesized radiolabeled insulin for each set of assays. Therefore, a rapid, non-radioactive, and reproducible assay is highly desirable.MethodsWe have developed electrochemiluminescence (ECL)-based assays that fulfill these criteria in the measurement of IAA and anti-insulin antibodies (IA) in non-obese diabetic (NOD) mice and in type 1 diabetic individuals, respectively. Using the murine IAA ECL assay, we examined the correlation between IAA, histopathological insulitis, and blood glucose in a cohort of female NOD mice from 4 up to 36 weeks of age. We developed a human IA ECL assay that we compared to conventional RBA and validated using samples from 34 diabetic and 59 non-diabetic individuals in three independent laboratories.ResultsOur ECL assays were rapid and sensitive with a broad dynamic range and low background. In the NOD mouse model, IAA levels measured by ECL were positively correlated with insulitis severity, and the values measured at 8-10 weeks of age were predictive of diabetes onset. Using human serum and plasma samples, our IA ECL assay yielded reproducible and accurate results with an average sensitivity of 84% at 95% specificity with no statistically significant difference between laboratories.ConclusionsThese novel, non-radioactive ECL-based assays should facilitate reliable and fast detection of antibodies to insulin and its precursors sera and plasma in a standardized manner between laboratories in both research and clinical settings. Our next step is to evaluate the human IA assay in the detection of IAA in prediabetic subjects or those at risk of type 1 diabetes and to develop similar assays for other autoantibodies that together are predictive for the diagnosis of this common disorder, in order to improve prediction and facilitate future therapeutic trials.

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Section: Resultssupporting

confidence: 72%

Section: Discussionmentioning

confidence: 99%

Antibodies against insulin measured by electrochemiluminescence predicts insulitis severity and disease onset in non-obese diabetic mice and can distinguish human type 1 diabetes status

Swafford

Shafer‐Weaver

et al. 2011

J Transl Med

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“…5,29,30 It may be higher in areas of low endemicity, with an antibody that is less specific to anti-PGL-1. In this context, a factor that can be investigated concerns the consumption of bovine and, as is well-known, Rio Grande do Sul consumes the most in Brazil.…”

Section: Discussionmentioning

confidence: 99%

Serology with ML Flow test in health professionals from three different states of Brazil

Calado

Magnanini

Moura

et al. 2013

An. Bras. Dermatol.

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BACKGROUND In highly endemic countries, transmission and sub-clinical infection of leprosy are likely and the disease manifests itself in individuals without any known close contact with a leprosy patient. Health workers are social contacts belonging to the same network (the Health System) and some of them share the same social environment (nursing assistants) as patients with known patients and / or carriers. OBJECTIVE To identify ML Flow seropositivity among health professionals. METHODS We conducted a cross-sectional study using a serological survey with the ML Flow test in 450 health professionals (doctors, nurses and nursing assistants), in order to detect seropositivity in areas of high and low endemicity in municipalities from three Brazilian states (RJ, MS and RS). RESULTS The results showed general 16% seropositivity, higher in low endemic areas, regardless of whether there was direct care for leprosy patients. Paradoxically, a statistical association was observed between the area studied and seropositivity, as the place with the lowest endemicity (CA) had the highest seropositivity rate (p = 0.033). CONCLUSION The authors suggest these results are associated with a presence of an unspecified link to bovine serum albumin (BSA), carrier of PGL-1 in the ML Flow test, and recommend expanded seroepidemiological research utilizing tests with human and bovine albumin.

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“…Anti-BSA antibodies present in human sera react with minor contaminants such as BSA in insulin preparations [7].…”

Section: Non-specific Reactions Involved In Indirect Elisamentioning

confidence: 99%

An ELISA protocol to improve the accuracy and reliability of serological antibody assays

Waritani¹,

Chang²,

McKinney³

et al. 2017

MethodsX

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Anti-BSA antibodies are a major cause of non-specific binding in insulin autoantibody radiobinding assays

Cited by 13 publications

References 10 publications

Antibodies against insulin measured by electrochemiluminescence predicts insulitis severity and disease onset in non-obese diabetic mice and can distinguish human type 1 diabetes status

Antibodies against insulin measured by electrochemiluminescence predicts insulitis severity and disease onset in non-obese diabetic mice and can distinguish human type 1 diabetes status

Serology with ML Flow test in health professionals from three different states of Brazil

An ELISA protocol to improve the accuracy and reliability of serological antibody assays

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