Immunoassay for quantifying squamous cell carcinoma antigen in serum.
BACKGROUND
Although the benefits of quantifying serum squamous cell carcinoma antigen (SCCa) have been reported, SCCa reagents were no longer available in the US by the late 1990s. Because SCCa quantification still has demonstrated clinical utility, we developed and validated a microtiter plate-based ELISA for measuring SCCa in serum.


METHODS
We coated microtiter strips overnight with capture anti-SCCa monoclonal antibody, washed the wells, added blocking buffer, and lyophilized the strips. For detection, we used a biotinylated anti-SCCa detection antibody, streptavidin/horseradish peroxidase conjugate, and tetramethylbenzidine/H(2)O(2) substrate. A novel blocking reagent against human antimouse antibodies (HAMA) was evaluated. A reference interval was established with sera from healthy individuals and was confirmed in smokers.


RESULTS
The assay was linear to 40 microg/L SCCa (slope, 1.00; y intercept, 0.695; R(2), 0.996) with a detection limit of 0.3 microg/L. The intraassay imprecision results [mean (CV)] were 2.5 microg/L (3.4%), 18.0 microg/L (3.0%), and 30.7 microg/L (2.4%); interassay imprecision results were 2.0 microg/L (9.9%), 20.0 microg/L (7.6%), and 36.3 microg/L (3.5%). A correlation analysis against an established automated assay generated a slope of 0.976 and a y intercept of -0.193 microg/L (r(2) = 0.916). An upper reference limit of 2.1 microg/L SCCa was established at 95% confidence level, with no difference observed in smokers. No correlation between SCCa concentration and age was observed (r(2) = 0.0003). At a blocking reagent concentration of 5 mg/L, HAMA interference was eliminated in 3 samples known to produce falsely increased SCCa results.


CONCLUSIONS
This SCCa ELISA demonstrates acceptable performance characteristics for quantifying serum SCCa and is effective in eliminating HAMA interference.
Erickson
J Alan
JA
ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, UT 84108, USA. ericksja@aruplab.com
Lu
Jun
J
Smith
Jeffery J
JJ
Bornhorst
Joshua A
JA
Grenache
David G
DG
Ashwood
Edward R
ER
D000328:Adult; D000368:Aged; D000951:Antigens, Neoplasm / Q000097:blood*; D014408:Biomarkers, Tumor / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D012016:Reference Values; D015843:Serpins / Q000097:blood*; D055815:Young Adult
D000951:Antigens, Neoplasm; D014408:Biomarkers, Tumor; D015843:Serpins; C051072:squamous cell carcinoma-related antigen