Development of glycan specific lectin based immunoassay for detection of prostate specific antigen

• Published in: International journal of biological macromolecules 2016-05, Vol.86, p.468-480
• Main Authors: Bhanushali, Paresh B., Badgujar, Shamkant B., Tripathi, Mukesh M., Gupta, Sanjeev, Murthy, Vedang, Krishnasastry, Musti V., Puri, Chander P.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Antibodies, Monoclonal - immunology; Biomarkers; Glycosylation; Humans; Immunoassay - methods; Lectin-based immunoassay; Lectins - metabolism; Polysaccharides - metabolism; Prostate cancer; Prostate specific antigen; Prostate-Specific Antigen - analysis; Prostate-Specific Antigen - blood; Prostate-Specific Antigen - chemistry; Prostate-Specific Antigen - immunology; Semen - metabolism; Substrate Specificity; Surface Plasmon Resonance
• ISSN: 0141-8130; 1879-0003
• DOI: 10.1016/j.ijbiomac.2016.01.110

[Display omitted] ⿢Prostate specific antigens (PSA) of seminal plasma and prostate cancer (PC) serum has been purified.⿢PSA of PC serum is a covalently complexed with α1-antichymotrypsin and protease activity of seminal PSA was discussed.⿢Characterization of a new anti-PSA monoclonal antibody was achieved for detection PSA in an immunoassay.⿢Fucosylated and sialylated linkages were analyzed in PSA of both origins (seminal and PC).⿢A lectin based immunoassay could be developed for the detection of changes in the glycosylation patterns of PSA of prostate cancer. We describe an analytical approach for the detection and verification of glycosylation patterns of prostate specific antigen (PSA), a key biomarker currently used for understanding the onset and prognosis of prostate cancer. PSA has been purified from the human seminal plasma and total PSA from prostate cancer sera. PSA is a monomeric glycoprotein with an apparent molecular mass 28040.467Da, which exhibits a characteristic protease activity against casein and gelatin. Its optimal protease activity is centered on neutral pH. Peptide mass fingerprint analysis of the purified PSA has yielded peptides that partially match with known database sequences (Uniprot ID P07288). Tryptic digestion profile of isolated PSA, infer the exclusive nature of PSA and may be additive molecule in the dictionary of seminal proteins. Surface plasmon resonance and lectin immunoassay revealed direct interaction between a newly developed anti-PSA monoclonal antibody (C4E6) and PSA. A lectin based immunoassay is reported here which was achieved with the C4E6 anti-PSA antibody and biotinylated plant lectins. This investigation provides an alternative method to isolate and quantify PSA with altered glycosylation which might be seen in the prostate cancer and developing a lectin based immunoassay to detect PSA in serum of prostate cancer patients.