Clinical Chemistry, Vol 39, 2154-2157, Copyright © 1993 by American Association for Clinical Chemistry

Competitive time-resolved immunofluorometric assay for quantifying carbonic anhydrase VI in saliva

S Parkkila, AK Parkkila, T Vierjoki, T Stahlberg and H Rajaniemi
University of Oulu, Department of Anatomy, Finland.

A competitive time-resolved immunofluorometric assay sensitive and robust enough for quantifying human salivary carbonic anhydrase isoenzyme VI (HCA VI) was developed. The solid-phase immunoassay is based on competition between Eu(3+)-labeled HCA VI and salivary HCA VI for polyclonal rabbit anti-HCA VI antibodies that are attached to microtiter plate wells precoated with sheep anti-rabbit IgG. The subsequent immunoassay including the separation of free and bound HCA VI requires only one incubation step, after which the Eu3+ of the bound labeled antigen is released into an enhancement solution. The highly fluorescent Eu chelates formed in this solution are then quantified by time-resolved fluorometry (Delfia). The time-resolution principle effectively obviates possible interferences from complex biological material such as saliva. The assay detection limit was 1.5 micrograms/L. Intra- and interassay imprecisions (CVs) were 5.1% and 5.3%, respectively. The mean analytical recovery was 93%. The mean +/- SD concentration of HCA VI in paraffin-stimulated saliva was 6.8 +/- 4.3 mg/L (n = 30) and the secretion rate was 10.2 +/- 7.9 micrograms/min. The method was useful for further investigations of the role of HCA VI in difficult matrices, e.g., saliva.

The following articles in journals at HighWire Press have cited this article:

				P. Karhumaa, J. Leinonen, S. Parkkila, K. Kaunisto, J. Tapanainen, and H. Rajaniemi The identification of secreted carbonic anhydrase VI as a constitutive glycoprotein of human and rat milk PNAS, September 5, 2001; (2001) 121172598. [Abstract] [Full Text] [PDF]

				J. Leinonen, S. Parkkila, K. Kaunisto, P. Koivunen, and H. Rajaniemi Secretion of Carbonic Anhydrase Isoenzyme VI (CA VI) from Human and Rat Lingual Serous von Ebner's Glands J. Histochem. Cytochem., May 1, 2001; 49(5): 657 - 662. [Abstract] [Full Text]

				P. Karhumaa, S. Parkkila, O. Tureci, A. Waheed, J.H. Grubb, G. Shah, A.-K. Parkkila, K. Kaunisto, J. Tapanainen, W.S. Sly, et al. Identification of carbonic anhydrase XII as the membrane isozyme expressed in the normal human endometrial epithelium Mol. Hum. Reprod., January 1, 2000; 6(1): 68 - 74. [Abstract] [Full Text] [PDF]

				J. Kivela, S. Parkkila, A.-K. Parkkila, J. Leinonen, and H. Rajaniemi Salivary carbonic anhydrase isoenzyme VI J. Physiol., October 15, 1999; 520(2): 315 - 320. [Abstract] [Full Text] [PDF]

				J. Saarnio, S. Parkkila, A.-K. Parkkila, A. Waheed, M. C. Casey, X. Y. Zhou, S. Pastoreková, J. Pastorek, T. Karttunen, K. Haukipuro, et al. Immunohistochemistry of Carbonic Anhydrase Isozyme IX (MN/CA IX) in Human Gut Reveals Polarized Expression in the Epithelial Cells with the Highest Proliferative Capacity J. Histochem. Cytochem., April 1, 1998; 46(4): 497 - 504. [Abstract] [Full Text]

				J. Kivela, S. Parkkila, A. Waheed, A.-K. Parkkila, W. S. Sly, and H. Rajaniemi Secretory carbonic anhydrase isoenzyme (CA VI) in human serum Clin. Chem., December 1, 1997; 43(12): 2318 - 2322. [Abstract] [Full Text] [PDF]

				P. Karhumaa, J. Leinonen, S. Parkkila, K. Kaunisto, J. Tapanainen, and H. Rajaniemi The identification of secreted carbonic anhydrase VI as a constitutive glycoprotein of human and rat milk PNAS, September 25, 2001; 98(20): 11604 - 11608. [Abstract] [Full Text] [PDF]