FILTER-PAPER BLOOD SPOT ASSAY OF HUMAN INSULIN-LIKE-GROWTH-FACTOR-I (IGF-I) AND IGF-BINDING PROTEIN-3 AND PRELIMINARY APPLICATION IN THE EVALUATION OF GROWTH-HORMONE STATUS

To facilitate broader applications of insulin-like growth factor I (IG F-I) and IGF-binding protein-3 (IGFBP-3) analysis, we developed proced ures for their measurements in extracts of whole blood dried on filter paper. A single 8-mm diameter filter paper disc containing about 13 m u L blood was used. IGFBP-3 was efficiently extracted in a buffer with in 1 h of incubation. IGF-I extraction involved incubation in buffer f ollowed by acidification and neutralization steps. Blood spot assays s howed intra- and interassay coefficients of variation (including inter spot variations) of 5.4-16.7% for IGF-I and 6.6-11.7% for IGFBP-3; rec overies were 97 +/- 7.1% and 101 +/- 8.7%, respectively. Recoveries of IGF-I and IGFBP-3 in response to 4- to 8-fold variations in extractio n buffer volume were 97 +/- 8.2% and 107 +/- 6.1%, respectively. Dried blood spot IGF-I and IGFBP-3 showed greater than 1-month stability at -20 C, 4 C, and room temperature and retained more than 65% of the im munoreactivity after approximately 1 month at 37 C. Both IGF-I and IGF BP-3 were contained within the plasma fraction of whole blood, and var iations (mean +/- SD) in IGF-I (204 +/- 29 mu g/L) and IGFBP-3 (4.4 +/ - 0.48 mg/L) measured in extracts of dried blood spot with adjusted he matocrit of 0.2-0.62 were acceptable. IGF-I and IGFBP-8 in paired plas ma and dried blood spot extracts of random samples (n = 46) showed exc ellent correlation (r > 0.94) with slopes of near unity. Compared to c onventional methods, the filter paper procedures were equally effectiv e in distinguishing IGF-I and IGFBP-3 levels in untreated GH receptor- deficient (n = 11) and age-matched normal controls (n = 16). We conclu de that blood collected on filter paper is ideal for IGF-I and IGFBP-3 analysis and may find applications in pediatric and large scale infan t screening programs.