Rat Multiple IgG2a/IgG1 Isotype Control [FITC+RPE]

Polyethylene glycol (PEG) represents an effective strategy to improve the pharmacokinetic profile of a molecule as it extends the biotherapeutic's half-life, masks immunogenic epitopes or modifies its distribution. The addition of one or multiple PEG moieties, in either linear or branched form, is known to carry the risk of potentially inducing an immunogenic response against PEG. The importance of accurately quantifying anti-PEG antibodies during a clinical study is well recognized and stems from the fact that anti-PEG antibodies have been shown to negatively impact the efficacy of the biotherapeutic that the PEG is coupled to. As a consequence, sponsors are encouraged to develop immunogenicity assays to assess appropriately the presence of anti-drug antibodies (ADA) against the protein component as well as the PEG. However, detection of anti-PEG antibodies is complicated by a number of technical challenges, including the availability of appropriate positive control material. In addition, the fact that some anti-PEG antibodies are known to circulate as low-affinity IgM, drives the need for an assay able to detect low affinity anti-PEG ADA even in the presence of high concentrations of the biotherapeutic. To address this need, we developed and validated an Affinity Capture Elution (ACE)-AGL assay to detect anti-drug and anti-PEG antibodies. In this assay, which we call ACE-AGL, ADA are captured by biotin-PEG-drug, acid eluted and re-captured on a second plate coated with protein AGL. ADA are then detected using Ruthenium-PEG-drug. The new assay format described is highly sensitive to both anti-drug and anti-PEG antibodies and very drug-tolerant. The ACE-AGL assay is easy to perform and has been successfully validated at two separate CROs. We propose the ACE-AGL format as a valid and effective alternative to the currently available assay methods.

Background Jamestown Canyon virus (JCV) is a mosquito-borne orthobunyavirus that causes acute febrile illness, meningitis, and meningoencephalitis, mainly among adults. JCV is widely distributed in North America and the number of JCV cases in the U.S. has increased in recent years. Therefore, the central nervous system disease caused by JCV can be considered a potentially re-emerging viral disease. However, the seroprevalence of JCV is unknown in Japan. The purpose of this study is to evaluate the seroprevalence of JCV in the Japanese population. Methods We used an IgG enzyme-linked immunosorbent assay (IgG-ELISA) with JCV-infected cell-lysates and/or a neutralizing (NT) antibody assay. The cut-off value of IgG-ELISA was determined using IgG-ELISA to analyze serum specimens from 37 healthy Japanese donors. IgG-ELISA was validated by assessing its sensitivity and specificity, using 38 human serum samples previously tested for the presence or absence of antibodies against JCV and snowshoe hare virus (SSHV), in an in-house NT antibody assay conducted by the Public Health Agency of Canada. The seroepidemiological study was performed using IgG-ELISA and NT antibody assay to analyze 246 human serum samples from the serum bank of the National Institute of Infectious Diseases (NIID) in Japan. Results The cut-off value of IgG-ELISA was determined at 0.20, based on the mean (- 0.075) and standard deviation (0.092) values using Japanese donors' sera. The sensitivity and the specificity of IgG-ELISA determined using 25 JCV-positive and 4 JCV-negative serum samples were 96 and 100%, respectively. Analysis of the 246 Japanese serum samples revealed that no specimen showed a higher value than the cut-off value of IgG-ELISA, and no sample tested positive by the NT antibody assay. Conclusions Our results showed that JCV is not circulating significantly in Japan. To the best of our knowledge, this is the first report to demonstrate the seroprevalence of JCV in the general population in Japan.