Specific ImmunoradiometricAssay of Insulin ... - Clinical Chemistry

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GL, Baxter RC, Bagley CL,. Wallace JC. Comparative binding ... J Immunol 1977;119:301-5. 29. Sevier ED, David GS, Martinis J, Desmond WJ, Bartholomew.
CLIN.CHEM.33/11, 2019-2023

(1987)

Specific ImmunoradiometricAssay of Insulin-likeGrowth Factor I with Use of Monoclonal Antibodies MItchell

G. Scott’ Gregory

C. Cuca,’ John R. Petersen,”4 Leon R. Lyle,’ B. Dan Burlelgh,2 and William H. Daughaday3

We identified two monoclonal antibodies that bind spatially

distinctepitopeson insulin-likegrowthfactor I (IGF-l). Using these two antibodies, we developed a simultaneous, two-site immunoradiometncassay (IRMA)specificfor IGF-l. This IRMA has no detectable cross reactivity with insulin, proinsulin, prolactin,or somatotropin,and less than 2% crossreactivity with IGF-lI. The assay response varies linearly with IGF-l concentrationsof 0-800 ig/L in serum;the detectionlimitis about 10 1zg/L.A comparisonof 26 IGF-l serumvalues from the IRMA and from a previouslyreported IGF-I specific RIA gave a correlationcoefficientof 0.96 with no substantialbias (slope = 1.10). IGF-l valuesfor serum,as an aid in assessing growth abnormalities,are easily (only three pipettingsteps) obtained in 99% of circulating IGF-I (9). Because secretion of IGF-I is regulated by somatotropin, this stability allows us to obtain information concerning chronic growth status from a random serum sample instead of necessitating dynamic or provocative somatotropin testing (10). With some exceptions, the concentrations of IGF-I in serum can be clinically important for assessing growth status (10-13). When used ‘Mallinckrodt, Inc., Diagnostic Products Division, 675 McDonnell Blvd., St. Louis, MO 63042. tmlnternational Minerals & Chemicals Corporation, Life Sciences Division, 1810 Frontage Road, Northbrook, IL 66044. tmWashington University School of Medicine, Metabolism Division, Box 8127, 660 South Euclid Ave., St. Louis, MO 63110. Current address: Invitron Corp., 4766 LaGuardia, St. Louis, MO 63134. 5Nonstandard abbreviations: IGF, insulin-like growth factor(s); MAb, monoclonal antibody(-iea); h, human origin; and ms,immunoradiometric assay. Received June 8, 1987; accepted August 24, 1987.

in coiijunction with somatotropin measurement, IGF-I concentrations can provide a valuable laboratory tool in differentiating the pathological conditions that produce growth abnormality (13). Nevertheless, IGF-I is not routinely measured in most clinical laboratories, owing to the presence of the IGFbinding protein complexes, which interfere with accuracy (13, 14). To eliminate this interference various extraction procedures-including acidification (15), acid-ethanol mixtures (14), acid gel filtration (7), and proteolytic cleavage (16)-have been tried. Non-immunological methods to measure IGF-I have included in vitro bioassays (17) and radioreceptor assays (18, 19), but these are impractical for routine use in clinical laboratories. The availability of specific polyclonal antisera (7), and, more recently, monoclenal antibodies (MAbs) (20-22), has permitted the development of liquid phase, competitive inhibition RIAs (7, 14-16, 20-22). Generally, however, these assays have been complicated by lengthy incubations, multiple steps, variable specificities, and tedious extraction procedures that usually necessitate neutralization or evaporation and reconstitution of the extracted samples (14). Here, we describe a unique set of MAbs, which we have used to developa monoclonal-antibody-based immunoradiometric assay (IRMA) for IGF-I. This assay has the advantages of short incubation, ease of performance, and