Avidin-biotin ELISA Research Articles

Measurement of free kappa immunoglobulin light chains in the cerebrospinal fluid by a competitive avidin—biotin ELISA

Measurement of free kappa immunoglobulin light chains in the cerebrospinal fluid by a competitive avidin—biotin ELISA

A general reagent for amplifying ELISAs

This paper describes a method to enhance the sensitivity of enzyme immunoassays by the use of solid tagged latex beads. Various bead sizes (0.21 and 0.81 μm) were tested using particles which had been covalently modified (or coated) with biotinylated rabbit anti-mouse immunoglobulin antibodies. Detection limits of mouse monoclonal antibodies or antigens (which have been reacted with specific mouse antibodies) were compared to those obtained using the general homogeneous ‘sandwich’ ELISA assay. Compared to the homogeneous biotin-avidin ELISA an increase in sensitivity level of about 6–10 times was observed an antigen detection assays. The stability of the reagent was excellent. The method gave low backgrounds and was as simple to use as the standard procedure.

Correlation between enterotoxigenicity, tested by different ELISA-techniques, antibiotic resistance patterns and phage groups of staphylococcus aureus strains

A group of 596 Staphylococcus aureus strains isolated from various clinical sources or implicated in food poisoning was investigated for enterotoxins A and B (SEA and SEB) production. The conventional ELISA techniques (competitive and sandwich ELISA) were compared with a newly developed avidin-biotin ELISA in their ability to detect the enterotoxins. The avidin-biotin system was not remarkably influenced by SPA up to 10 μg/ml. A semi-quantitative competitive ELISA for the detection of staphylococcal protein A (SPA) in culture supernatants was carried out in parallel. The strains isolated in cases of food poisoning showed different antibiotic resistance patterns, whereas the strains from clinical sources were selected for either methicillin or penicillin resistance only. The strains isolated in food poisoning outbreaks (FP strains) were enterotoxin A positive in 22%, enterotoxin B positive in 11%, and SEA + SEB positive in 9% of cases. The strains with resistance to penicillin only (PE R strains) produced SEB in 26%, SEA in 14%, and both toxins in 7% of the cases. The methicillin-resistant strains (MC R strains) produced SEA in 59% of cases, whereas SEB was produced in 6% only (SEA + SEB: 20%). 37% of the SEA producers belonged to phage group III (SEB: 30%; SEA + SEB: 25%) and 12% (SEB: 11%; SEA + SEB: 9%) to phage group I. 26% of the SEA-producing and 37% of the SEB-producing strains (SEA + SEB: 23%) were non-typable.

Effect of praziquantel on certain immune responses of schistosomal Egyptian patients. I. Changes of specific immunoglobulins.

Specific antischistosomal IgG, IgM, and IgE were estimated by ELISA in 117 rural school students before specific treatment with praziquantel monthly for 3-4 months thereafter. IgG and IgM were estimated as percentage of bound antibodies. IgE was estimated by avidin-biotin ELISA (AB-ELISA) as IU/ml using a panel of known IgE standards. Soluble surface Schistosoma mansoni adult worm antigen was used for all estimates. Total IgE was estimated in a smaller group by an ELISA kit. The percentage of specific IgE was calculated. A group of endemic controls (22 students) and non-endemic controls (17 cases) were included. Statistical analysis of results showed the specific immunoglobulins to be significantly reduced 2 months after treatment of the schistosomal cases. These reduced levels, however, were still significantly higher than those of controls. The presence of early hepatosplenomegaly and the co-existence of other parasites had no significant influence on the results. No correlation could be established between the levels of specific antischistosomal IgG, M and E and the intensity of infection. The significance of these results is discussed.

An avidin-biotin ELISA assay for the measurement of de novo human IgE synthesis in culture supernatants

A very sensitive (100 pg/ml) solid-phase enzyme immunoassay (ELISA) for the determination of human IgE has been developed. This assay incorporates the avidin-biotin system to increase sensitivity and can detect as little as 100 pg/ml (10 pg/test) of human IgE. The assay is highly specific and allows quantitative determination of human IgE in supernatants of peripheral blood lymphocytes as well as in serum. The very high sensitivity of the assay was accomplished by optimizing concentrations of the following reagents: (1) affinity-purified rabbit anti-human IgE coating antibodies; (2) biotin-conjugated goat anti-human IgE; (3) avidin-horseradish peroxidase (HRP) conjugate. In summary, the assay described is rapid (6 h), reproducible, isotype specific, and has the sensitivity of radioimmunoassays usually employed for the quantification of IgE. This assay may be utilized in establishing concentrations of in vitro IgE levels synthesized by human peripheral blood lymphocyte (PBL).

Monoclonal Antibody-based Biotin-Avidin ELISA for the Detection of Soybean Mosaic Virus in Soybean Seeds

SUMMARY Two monoclonal antibodies (M-Ab) specific for different epitopes on particles of soybean mosaic virus (SMV) were used in a double-antibody sandwich ELISA (M-Ab ELISA). The non-isotopic immunoassay, which used a biotinylated second antibody and an avidin-alkaline phosphatase detection system to detect SMV in soybean seed extracts, was compared with a polyclonal antibody-based solid-phase radioimmunoassay (SPRIA). m-Ab ELISA detected less than 10 ng of SMV/ml and was more sensitive than the SPRIA which detected 25 ng SMV/ml. Furthermore, m-Ab ELISA required less than 36 h for seed sample assays and only 5.5 h for assays involving purified virus, whereas SPRIA required 3 or 4 days. When seeds from 33 field plots, in which 0%, 30% and 50% of the soybean plants had been inoculated with SMV, were assayed by both systems, results of the two tests correlated for 31 of 33 (94%) seed samples. This suggests that dual-site biotin-avidin m-Ab ELISA systems have potential utility for routine screening of seed samples.

An avidin-biotin ELISA for the measurement of serum and secretory IgD

This paper described an improved microtiter solid-phase enzyme immunoassay for the determination of serum and secretory IgD. Use of the interaction between biotinylated anti-human IgD and horseradish peroxidase(HRP)-avidin conjugate permits quantitation of human IgD in the range of 1–64 ng/ml. IgD was detected in all samples of serum, saliva and nasal secretions of 28 normal adults. In only one subject both serum and secretory IgD were undetectable. The mean concentration of serum IgD determined by this assay is similar to that reported by other authors using radioimmunoassay. The assay described is not only rapid and inexpensive but at least as sensitive as the radioimmunoassays usually employed for quantitation of IgD.