Endogenous protein interference

In general, bioassays have more technical issues due to serum effects on cells, greater variability and sensitivity. Additionally, both bioassays and CLB assays are complicated by interferences from dmg and/or endogenous proteins that may be difficult to overcome. 

Tetrazolium salts can be reduced nonselectively by many endogenous reductants such as thiol-containing proteins, as well as exogenous ones such as ascorbic acid.521 -523 This can lead to serious interferences and several measures have been described to reduce or eliminate their effect.524,525,650... 

Of particular interest is the comparison of the performance of cation exchange and mixed mode sorbents for their efficacy in cleaning up endogenous phospholipids. Unlike the protein-related materials that are eluted in the very early stages of HPLC, these phospholipids elute in the hydro-phobic region and interfere with drug peaks which also elute around the same time. 

Since FPIAs are conducted as homogeneous immunoassays, they are susceptible to effects from endogenous fluorophores and from intersample variations. Such problems and others due to the sample matrix are largely avoided by sample dilutions of several hundredfold. Low-affinity, nonspecific binding of tracers to sample proteins, when present in sufficiently high concentrations, can result in a falsely elevated polarization signal. Interference from sample proteins can be eliminated when warranted, by proteolytic hydrolysis with pepsin.(46)... 

The primary objective of integration plot analysis is to analyze the data on influx of the test substrate from the circulating blood to the retina (i.e., blood-to-retina direction) across the BRB after intravenous administration of the test substrate. The advantage of this approach is that it allows reliable determination of the retinal uptake (i.e., clearance) of the test substrate which has a slow permeability across the BRB [28], On the other hand, due to the intravenous injection, interference by endogenous substrates and plasma-protein binding of the test substrate can produce an unseemingly low estimate of the retinal uptake. 

---