The aim of the present study was to evaluate the suitability of dark hair as a matrix for determination of the beta(2)-adrenergic agonist clenbuterol residues using previously validated enzyme-linked immunosorbent assay (ELISA) as a screening method for its quantitative determination. The experimental group of mice (n = 60) were treated with two different anabolic dosages of clenbuterol for 15 days, whereas the control group of animals (n = 30) was left completely untreated. Hair samples were collected on days 0, 5, 10, and 15 of treatment. Validation of the ELISA analytical procedure showed good recovery (mean recovery 74%) with an acceptable intra-assay variation in individual measurements for all hair samples to which 5, 10, and 50 ng/g clenbuterol were added (CV < 10%). Low blank levels of clenbuterol (2.4 +/- 0.6 ng/g) were measured in hair of untreated mice, whereas significantly higher clenbuterol concentrations rising proportionally with the time of treatment were recorded in hair of mice treated with lower (6.5 mg/kg body mass) and higher (12.5 mg/kg body mass) dose of clenbuterol. The peak hair concentration of clenbuterol measured on the last day of treatment (day 15) was 1553.9 +/- 140.1 ng/g and 6248.3 +/- 589.4 ng/g in the lower and higher dose group, respectively. Study results clearly indicated dark hair as a pigmented tissue to have a high accumulation potential for clenbuterol residues, thus being the target matrix of choice for detection of clenbuterol abuse as an anabolic in meat production.