The use of antibiotics grown promoter has been banned, due led problems with drug residues in animal products and increased bacterial resistance. For several reasons there is a growing interest in the scientific community in immunoglobulin Y as antibiotic alternative and their oral administration in the polyclonal antibody (pAb) format, to maintain animal health and performance, do not require IgY purification for large-scale production, resulting in protein impurities and high concentration of biotin in the samples. The signal amplification through non-covalent interaction of biotin with streptavidin has been extensionally used by many laboratories for several diagnosis diseases and scientific research in enzyme-linked immunosorbent assay (ELISA). However, the riches sources of biotin as egg yolk, leading to high biotin concentrations in samples, harming the accuracy of diagnostic and proteins concentrations tests. This study aimed to evaluate the biotin influence on measurement of Immunoglobulin Y in egg yolk freeze dried samples from immunized laying hens by immunoassays using biotin-avidin/streptavidin. The IgY concentration changed down in immunoassays using biotin-avidin/streptavidin. The detection of IgY in yolk samples by ELISA using streptavidin–biotin binding as part of the assay methodology requires some technology to neutralize high concentration of biotin on sample or more steps beyond delipidation to isolate the target protein. Otherwise, an ELISA without the use of streptavidin-biotin binding would be more advisable to avoid biotin and targets protein relationship and prevent biotin interference on results.

Background: The duration of the vaccine's protective efficacy against SARS-CoV-2 is unknown. Evaluation of the clinical performance of available tests is required. Objectives: To evaluate the clinical performance of three immunoassays for the detection of IgG antibodies generated by mRNA vaccines against SARS-CoV-2. Methods: Two automated immunoassays (Euroimmun Anti-SARS-CoV-2 ELISA IgG and Abbott SARS-CoV-2 CLIA IgG) and one lateral flow immunoassay (LFIA Test Livzon IgG) were tested. 300 samples distributed in 3 groups were analyzed: 100 subjects over 18 years old and under 45 years old, 100 subjects between 45-65 years old and another 100 over 65 years old. collected before vaccination, at 21 days, 1, 2, 3 and 6 months post-vaccination. Sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, negative likelihood ratio, and agreement (I. Kappa) were calculated for each serological test. Results: Maximum sensitivity for IgG was 98.7%, 98.1%, and 97.8% for the ELISA Euroimmun, CLIA Abbott, and Livzon LFIA assays and maximum specificity for IgG was 99.4%, 99.9%. % and 98.4% ELISA, CLIA and LFIA respectively at 3 months after vaccination with a decrease in antibody levels from the sixth month. The best agreement was observed between ELISA and CLIA 100%; (k = 1.00). The agreement between ELIA, CLIA and LIFIA was 99% (k = 0964) at the second and third month after vaccination. Seroconversion was faster and longer lasting in the younger age groups. Conclusion: Our study showed an equivalent and homogeneous clinical performance for IgG of three immunoassays after vaccination and that the LIFIA assay is the most cost-effective, reliable and accurate for routine use in population studies of seroconversion and seroprevalence.

The primary screening of hybridoma cells is a time-critical and laborious step during the development of monoclonal antibodies. Often critical errors occur in this phase, which supports the notion that the generation of monoclonal antibodies with hybridoma technology is difficult to control and hence a risky venture. We think that it is crucial to improve the screening process to eliminate most of the immanent deficits of the conventional approach. With this new microarray-based procedure, several advances could be achieved: Selectivity for excellent binders, high throughput, reproducible signals, avoidance of misleading avidity (multivalency) effects, and simultaneous performance of competition experiments. The latter can directly be used to select clones of desired cross-reactivity properties. In this paper, a model system with two excellent clones against carbamazepine, two weak clones and blank supernatant has been designed to examine the effectiveness of the new system. The excellent clones could be detected largely independent of the IgG concentration, which is unknown during the clone screening since the determination and subsequent adjustment of the antibody concentration is not possible in most cases. Furthermore, in this approach, the enrichment, isolation, and purification of IgG for characterization is not necessary. Raw cell culture supernatant can be used directly, even when fetal calf serum (FCS) or other complex media had been used. In addition, an improved method for the oriented antibody-immobilization on epoxy-silanized slides is presented. Based on the results of this model system, we conclude that this approach should be preferable to most other protocols leading to many of false positives, causing expensive and lengthy confirmation steps to weed out the poor clones.

Thousands of antibodies for diagnostic and other analytical purposes are on the market. However, it is often difficult to identify duplicates, reagent changes, and to assign the correct original publications to an antibody. This slows down scientific progress and might even be a cause of irreproducible research and a waste of resources. Recently, activities were started to suggest the sole use of recombinant antibodies in combination with the open communication of their sequence. In this case, such uncertainties should be eliminated. Unfortunately, this approach seems to be rather a long-term vision since the development and manufacturing of recombinant antibodies remain quite expensive in the foreseeable future. Also, nearly all commercial antibody suppliers may be reluctant to publish the sequence of their antibodies, since they fear counterfeiting. De-novo sequencing of antibodies is also not feasible today for a reagent user without access to the hybridoma clone. Nevertheless, it seems to be crucial for any scientist to have the opportunity to identify an antibody undoubtedly to guarantee the traceability of any research activity using antibodies from a third party as a tool. For this purpose, we developed a method for the identification of antibodies based on a MALDI-TOF-MS fingerprint. To circumvent lengthy denaturation, reduction, alkylation, and enzymatic digestion steps, the fragmentation was performed with a simple formic acid hydrolysis step. Eighty-nine unknown monoclonal antibodies were used for this study to examine the feasibility of this approach. Although the molecular assignment of peaks was rarely possible, antibodies could be easily recognized in a blinded test, simply from their mass-spectral fingerprint. A general protocol is given, which could be used without any optimization to generate fingerprints for a database. We want to propose that in most scientific projects relying critically on antibody reagents, such a fingerprint should be established to prove and document the identity of the used antibodies and to assign a specific reagent to a datasheet of a commercial supplier, a public database record or an antibody ID.