Isoelectric focusing of group specific component in semen and vaginal stains on ultrathin immobilized pH gradient gels

Abstract

Subtyping of the group specific component in secretions of semen and vaginal fluids is impossible with conventional detection systems. By means of a highly sensitive alkaline-phosphatase secondary antibody system the group specific component can reliably be detected in semen stains. Results with vaginal swabs were inconsistent and in saliva stains Gc activity could not be detected.

Water analysis

“…Gc subtypes have been identified by isoelectric focusing (355)(356)(357)(358)(359). Isoelectric focusing of Gc was reported for semen, blood, and vaginal fluid (396,397). Isoelectric focusing methods for Hp typing have been described (360,361), as well as a micromethod for phenotyping Hp in bloodstains (362).…”

Group‐specific component subtyping in semen and seminal stains by conventional isoelectric focusing

“…Using isoelectric focusing on ultrathin immobilized pH gradient gel, Pflug etaf. [3], Pflug [4] and Potsch-Schneider and Klein [5] demonstrated GC subtypes also in semen and seminal stains. We have recently observed that much better isoelectric focusing patterns of GC in human semen can be obtained by treating the samples with neuraminidase.…”

Immobilized pH gradients

“…Moreover, the contact of the sticky IPG surface with nitrocellulose membranes is better avoided, pouring an agarose coating 58 or overlaying a cellulose acetate foil 60. As an alternative, diffusion blotting may be used 56, 61. An overnight transfer, in the presence of 4 M guanidinium chloride, seems to provide high protein recovery even after TCA/SSA precipitation in provision for detergent elution; 4 M guanidinium chloride also prevents IPG from sticking to PVDF membranes 56.…”

“…It is less surprising not to find in recent years any report dealing with clinical biochemistry applications of IPGs. Setup for resolution of human serum proteins that used to be phenotyped by IEF dates back to the '80s: group specific component 61, 73, transferrin 74 and mostly α 1 ‐antitrypsin 74–77. In the meantime, most of the clinical biochemistry and forensic medicine typing has switched from phenotype to genotype, taking advantage of the acknowledged features of DNA – stoichiometric presence in all cell types from embryo to adult stage, chemical stability, easy amplification.…”