Parallelism Experiments to Evaluate Matrix Effects, Selectivity and Sensitivity in Ligand-Binding Assay Method Development: Pros and Cons

Tu, Jing; Bennett, Patrick

doi:10.4155/bio-2017-0084

Cited by 45 publications

(16 citation statements)

References 42 publications

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“…For ligand binding assays (e.g., immunoassays), two key properties are specificity and selectivity 28 . The assay specificity refers to how well the assay (with its antibodies and other components) can distinguish between its intended analyte and other structurally similar components.…”

Section: Factors Affecting Reproducibilitymentioning

confidence: 99%

Increasing the reproducibility of fluid biomarker studies in neurodegenerative studies

et al. 2020

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Biomarkers have revolutionized scientific research on neurodegenerative diseases, in particular Alzheimer’s disease, transformed drug trial design, and are also increasingly improving patient management in clinical practice. A few key cerebrospinal fluid biomarkers have been robustly associated with neurodegenerative diseases. Several novel biomarkers are very promising, especially blood-based markers. However, many biomarker findings have had low reproducibility despite initial promising results. In this perspective, we identify possible sources for low reproducibility of studies on fluid biomarkers for neurodegenerative diseases, with a focus on Alzheimer’s disease. We suggest guidelines for researchers and journal editors, with the aim to improve reproducibility of findings.

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Section: Factors Affecting Reproducibilitymentioning

confidence: 99%

Increasing the reproducibility of fluid biomarker studies in neurodegenerative studies

et al. 2020

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“…We also evaluated the peptide-level similarity between Mus and Acomys for each gene represented on the array. Parallelism was examined using standard protocols (45,46). Briefly, samples from species and source were randomly pooled to provide a representative cytokine concentration and were run in triplicate at serial dilutions (1:2, 1:6, 1:18, 1:54, and 1:162).…”

Section: Cytokine Assaymentioning

confidence: 99%

Complex Tissue Regeneration in Mammals Is Associated With Reduced Inflammatory Cytokines and an Influx of T Cells

et al. 2020

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While mammals tend to repair injuries, other adult vertebrates like salamanders and fish regenerate damaged tissue. One prominent hypothesis offered to explain an inability to regenerate complex tissue in mammals is a bias during healing toward strong adaptive immunity and inflammatory responses. Here we directly test this hypothesis by characterizing part of the immune response during regeneration in spiny mice (Acomys cahirinus and Acomys percivali) vs. fibrotic repair in Mus musculus. By directly quantifying cytokines during tissue healing, we found that fibrotic repair was associated with a greater release of pro-inflammatory cytokines (i.e., IL-6, CCL2, and CXCL1) during acute inflammation in the wound microenvironment. However, reducing inflammation via COX-2 inhibition was not sufficient to reduce fibrosis or induce a regenerative response, suggesting that inflammatory strength does not control how an injury heals. Although regeneration was associated with lower concentrations of many inflammatory markers, we measured a comparatively larger influx of T cells into regenerating ear tissue and detected a local increase in the T cell associated cytokines IL-12 and IL-17 during the proliferative phase of regeneration. Taken together, our data demonstrate that a strong adaptive immune response is not antagonistic to regeneration and that other mechanisms likely explain the distribution of regenerative ability in vertebrates.

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“…We validated the IgA assay by checking for parallelism, accuracy, and intra‐ and inter‐assay coefficient of variation (CV). Testing for parallelism is important when commercial kits are used with fecal samples as these kits are designed for use with serum, and endogenous compounds in the fecal matrix may interfere with the binding of the IgA to the antibody (Tu & Bennett, 2017). Second, in a small set of samples, we assessed experimentally whether sample collection and storage conditions in the field affect IgA concentrations.…”

Section: Introductionmentioning

confidence: 99%

Noninvasive measurement of mucosal immunity in a free‐ranging baboon population

Gesquiere

Habig

Hansen

et al. 2020

American J Primatol

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Ecoimmunological patterns and processes remain understudied in wild primates, in part because of the lack of noninvasive methods to measure immunity. Secretory immunoglobulin A (sIgA) is the most abundant antibody present at mammalian mucosal surfaces and provides an important first line of defense against pathogens. Recent studies show that sIgA can be measured noninvasively in feces and is a good marker of mucosal immunity. Here we validated a commercial ELISA kit to measure fecal IgA in baboons, tested the robustness of its results to variation in collection and storage conditions, and developed a cost‐effective in‐house ELISA for baboon fecal IgA. Using data from the custom ELISA, we assessed the relationship between fecal IgA concentrations and gastrointestinal parasite burden, and tested how sex, age, and reproductive effort predict fecal IgA in wild baboons. We find that IgA concentrations can be measured in baboon feces using an in‐house ELISA and are highly correlated to the values obtained with a commercial kit. Fecal IgA concentrations are stable when extracts are stored for up to 22 months at −20°C. Fecal IgA concentrations were negatively correlated with parasite egg counts (Trichuris trichiura), but not parasite richness. Fecal IgA did not vary between the sexes, but for males, concentrations were higher in adults versus adolescents. Lactating females had significantly lower fecal IgA than pregnant females, but neither pregnant nor lactating female concentrations differed significantly from cycling females. Males who engaged in more mate‐guarding exhibited similar IgA concentrations to those who engaged in little mate‐guarding. These patterns may reflect the low energetic costs of mucosal immunity, or the complex dependence of IgA excretion on individual condition. Adding a noninvasive measure of mucosal immunity will promote a better understanding of how ecology modulates possible tradeoffs between the immune system and other energetically costly processes in the wild.

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Parallelism Experiments to Evaluate Matrix Effects, Selectivity and Sensitivity in Ligand-Binding Assay Method Development: Pros and Cons

Cited by 45 publications

References 42 publications

Increasing the reproducibility of fluid biomarker studies in neurodegenerative studies

Increasing the reproducibility of fluid biomarker studies in neurodegenerative studies

Complex Tissue Regeneration in Mammals Is Associated With Reduced Inflammatory Cytokines and an Influx of T Cells

Noninvasive measurement of mucosal immunity in a free‐ranging baboon population

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