Abstract
Objective Glypican4 is an interesting new adipokine, which
seems to play an important role in developmental processes and is
potentially associated with metabolic changes in obesity and type 2 diabetes
mellitus. Currently, only a few studies examined glypican4 in human blood,
mainly in adults.
Design, Patients and Measurements The aim of our study was to
investigate glypican4 serum levels in lean, overweight, and obese children
and adolescents, to unravel a possible association between glypican4 serum
levels and parameters of obesity and insulin resistance. In order to
determine a suitable method for investigating glypican4 serum levels, we
validated two commercially available human glypican4 ELISA kits, using serum
and plasma samples of an obese, insulin-resistant patient, and a healthy
control subject, a human recombinant glypican4 protein fragment and
glypican4-overexpressing cell lysate.
Results Using ELISA kit #1 we were not able to detect values
above background level, apart from standard curve values. ELISA kit #2
initially seemed suitable to measure glypican4, but further validation
experiments showed non-linearity of serial dilutions, no recognition of a
human recombinant glypican4 protein fragment and non-linearity in the
recovery of glypican4-overexpressing cell lysate. In addition, there was a
considerable decrease (approx. 68%) of measured values between two
experiments, performed at different time points with aliquots of the same
serum sample. Contrary to that, further experiments found sample stability
not to be compromised.
Conclusions Extensive evaluation of the performance of two
commercially available ELISA kits led to the conclusion that none of them is
applicable for the measurement of glypican4 in human blood samples.
Key words
adipocytokines - obesity - humans - insulin resistance - obesity - enzyme-linked immunosorbent assay - diabetes - autoimmunity
