Epitope characterization can also help elucidate the mechanism of antibody binding (Davidson et al. The identification and characterization of antibody binding sites aid in the discovery and development of new therapeutics, vaccines, and diagnostics (Gershoni et al. 1991).Įpitope mapping is the process of experimentally identifying the binding site, or ‘epitope,’ of an antibody on its target antigen (usually a protein) (Dejana and Corada 1999 Westwood and Hay 2001 Davidson and Doranz 2014). In addition, rpS3 is overexpressed in colorectal cancer cells, which suggests that there may be a relationship between the occurrence of colorectal cancers and rpS3 (Pogue-Geile et al. Therefore, rpS3 can cleave the phosphodiester bonds at the pyrimidine dimer generated by UV irradiation. In fact, rpS3 appears to have a wide range base-damage-endonuclease activity which recognizes various kinds of damaged lesions, including AP sites, thymine glycols and pyrimidine dimers (Kim et al. The AP endonuclease activity of rpS3 functions as a lyase through β-elimination. Also, rpS3 appear to aid damaged DNA processing by cleaving apurinic/apyrimidic (AP) sites that is, when DNA is damaged, the base of the damaged area is cut off with the function of another DNA glycosylase to form an AP site, and then the AP site is cleaved by an AP endonuclease activity of rpS3. Furthermore, it was recently discovered that the rpS3 protein associates with the TFIIH complex and positively regulates UV-damaged nucleotide excision repair by assisting XPD helicase and increasing the turnover rate of TFIIH complex (Park et al. The rpS3 protein has extra-ribosomal functions it is first discovered to be UV endonuclease III, which functions as an endonuclease to repair DNA damage caused by ultraviolet light (UV) (Kim and Linn 1989 Kim et al. Also, rpS3 is reported to be involved in the regulation of its own mRNA transcription (Polakiewicz et al. The human RPS3 gene is located on 11q13.3-q13.5 on human chromosome 11. RpS3 is located on the outer surface of the 40S subunit of the ribosome and plays an important role in protein synthesis in vivo (Westermann et al.
RpS3 has a mass of 26.69 kDa and an amino acid length of 243. Ribosomal protein S3 (rpS3), also known as ribosomal protein uS3, is a member of 40S small ribosomal subunit encoded by the RPS3 gene. Ribosomes, the protein synthesis machinery of all living cells, are composed of several rRNAs and ribosomal proteins. coli using pET-15b vector.įor cells to survive, proteins must be continuously synthesized.
This study relates to a mAb of human rpS3 (ribosomal protein small subunit 3, ), particularly, a mAb produced by using recombinant protein expressed and purified with E. In addition, useful screening results must be obtained relatively independent of the mAb concentration in the supernatants because optimization of the ELISA parameters (such as supernatant dilution and coating conjugate dilution) prior to the screening is usually time-consuming or even impossible, especially as the screening often involves a single measurement per mAb. The screening method should clearly detect high-affinity mAbs with a minimum of both false positives and false negatives. Therefore, an ideal screening method should be fast, reliable, and easy to perform, especially if the laboratory does not have an equipment for performing automated immunoassays. Irrespective of the intended application, selection of high-affinity mAbs were often preferred, and an efficient hybridoma screening procedure is a critical step that usually must be completed in a short time (Burrin and Newman 1991 Jang et al.
Immunization of animals, mainly mice, in combination with hybridoma technology, is still the most common method for generating mAbs. In addition, antibodies are increasingly used as human therapeutics. Monoclonal antibodies (mAbs) are useful biological tools for various analytical applications, including clinical chemistry, food analysis, and environmental monitoring.
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