E. Testai - Istituto Superiore di Sanità
E. Testai - Istituto Superiore di Sanità
E. Testai - Istituto Superiore di Sanità
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METODI PER DETERMINARE LA PRESENZA DI CYP E LORO<br />
ATTIVITA’ NEGLI ORGANISMI IN VIA DI SVILUPPO<br />
A) Detection of specific enzymatic activities with model (probe)<br />
substrates (Substances which are specifically metabolized by one CYP<br />
lea<strong>di</strong>ng to a easily detectable product)<br />
CYP1A1<br />
Ethoxyresorufin Resorufin (Fluorescent)<br />
CYP3A4<br />
Eritrhromycin Demethylated product + HCHO<br />
LIMITS: Low activities (below limit of detection) Overlapping in substrate<br />
specificity, Probe substrates suitable for adult CYPs.<br />
A) Enzyme localization by means of immunoblotting or immunohysto-<br />
chemical tecniques<br />
LIMITS: Cross-reactivity of antibo<strong>di</strong>es towards rat/mouse CYP(s) with more<br />
than one human CYP.<br />
A) Use of PCR and reverse-transcriptase (RT)-PCR to detect even small<br />
quantities of specific mRNA<br />
ADVANTAGES: increase in sensitivity and specificity.
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