Comparison of two in vitro methods for the measurement of recombinant human TSH bioactivity.
Thyroid stimulating hormone (TSH), a pituitary glycoprotein hormone, is a potent inducer of intracellular cAMP production. Two methods for measuring TSH bioactivity were evaluated and compared. One assay is based on using a radioimmunoassay (RIA) to measure the recombinant human TSH-induced increase in cAMP using a bovine thyroid membrane isolate. The other is based on a Chinese hamster ovary (CHO) cell line that has been transfected with the TSH receptor and a cAMP-responsive luciferase reporter. The within-assay coefficient of variation for the membrane-based assay was determined to be approximately 35% compared with approximately 25% for the cell-based assay. Twenty-one preparations of recombinant human TSH (rhTSH) were tested using both methods. No significant difference was detected between the data sets and no assay bias was present. Both assay systems provide a suitable means for measuring the activity of rhTSH. The advantage of the membrane-based assay is the relatively small quantity of TSH needed for analysis. However, the average time required to analyse a sample using the membrane-based method was more than twice as long as that needed to test a sample in the cell-based assay. Other advantages of the cell-based method include the use of a 96-well format, which facilitates the analysis of several concentrations of rhTSH within one assay plate, and the use of a non-radioactive endpoint.
Sendak
Rebecca A
RA
Therapeutic Protein Research and Development, Genzyme Corporation, One Mountain Road, Framingham, MA 01701, USA. rebecca.sendak@genzyme.com
Wang
Fei
F
Geagan
Laura B
LB
Armstrong
Lori A
LA
Thyne
Charles D
CD
Cole
Edward S
ES
Mattaliano
Robert J
RJ
D000818:Animals; D001681:Biological Assay / Q000379:methods*; D016466:CHO Cells; D002417:Cattle; D006224:Cricetinae; D017930:Genes, Reporter; D006801:Humans; D066298:In Vitro Techniques; D008156:Luciferases / Q000235:genetics; D011863:Radioimmunoassay / Q000379:methods; D011989:Receptors, Thyrotropin / Q000235:genetics; D011994:Recombinant Proteins / Q000032:analysis / Q000494:pharmacology; D013961:Thyroid Gland / Q000187:drug effects; D013972:Thyrotropin / Q000032:analysis* / Q000494:pharmacology*; D014162:Transfection
D011989:Receptors, Thyrotropin; D011994:Recombinant Proteins; D013972:Thyrotropin; D008156:Luciferases