EVALUATION OF A RADIOIMMUNOASSAY FOR DETERMINATION OF CALCITRIOL IN HUMAN SERA EMPLOYING A I-125 LABELED TRACER

The performance characteristics of a radioimmunoassay employing a I-12 5-labelled tracer for determination of calcitriol in human sera are re ported. The assay is based on an immunoextraction step employing a mon oclonal antibody against calcitriol followed by a radioimmunoassay (us ing I-125-labelled calcitriol and sheep antiserum against calcitriol). Bound/free separation is performed with an anti-sheep IgG antibody bo und to cellulose. Within-run imprecision (n = 20) was 12.8% (mean = 9. 4 ng/l) and 11.1% (mean = 48.8 ng/l), between-day imprecision (n = 11) was 27.1% (mean = 9.6 ng/l) and 17.2% (mean = 47.2 ng/l). Linearity o f dilution was investigated by mixing pooled sera containing 62.0 ng/l and 4.7 ng/l calcitriol, respectively. The relationship between measu red and expected concentrations was characterized by a linear correlat ion coefficient of r = + 0.990. The values obtained with the I-125-bas ed radioimmunoassay were compared with those obtained with a radiorece ptor-assay using a H-3-labelled tracer; the regression line was y = 1. 091 x - 4.545 (n = 84; r = + 0.935), where y = calcitriol [I-125] [ng/ l] and x = calcitriol [H-3] [ng/l]. Mixing 9 volumes of sera from pati ents with renal insufficiency (n = 7) with 1 volume of 'calibrator F' (assigned value: 227 ng/l) yielded recovery rates of 90 +/- 8% (mean /- SD). The detection limit was 3.0 ng/l. The cross-reactivity of chol ecalciferol metabolites was found to be < 0.00003 for 25-hydroxycholec alciferol, 24R,25-dihydroxycholecalciferol and 25S,26-dihydroxycholeca lciferol. A preliminary reference interval (5th to 95th percentile) wa s established in 40 apparently healthy persons (17 males and 23 female s; age range: 20-61 [mean: 32] years) (19-74 ng/l). The method present ed shows high practicability and may therefore be considered as a usef ul alternative to cumbersome assays using H-3-labelled tracers.