ABSTRACT
BACKGROUND: The recent elucidation of the importance of serological free prostate-specific antigen (PSA) in the diagnosis of prostate cancer has created a demand for immunoassays specific for free PSA. METHODS: We developed and characterized 11 monoclonal antibodies with high affinities for PSA (Ka values from 1.1 x 10(8) to 1.8 x 10(10)L/mol), only 3 of which cross-react with human glandular kallikrein (hK2). Using these antibodies and PSA antibodies developed by others, in conjunction with time-resolved fluorometry, we developed ultrasensitive sandwich immunoassays specific for the free form of PSA. RESULTS: The analytical detection limit of these immunoassays is 0.001 microg/L. To our knowledge, this is the most sensitive free PSA assay reported to date. The free PSA immunoassays exhibit <1% cross-reactivity with PSA-alpha1-antichymotrypsin, show no cross-reactivity with hK2, and correlate well with established free PSA kits. The 11 antibodies developed by our group, in conjunction with 4 commercially available antibodies, were used to generate a putative epitope map of the PSA molecule. CONCLUSION: The highly sensitive free PSA immunoassays may be used for measuring PSA subfractions in female serum, an application currently impossible with other reported free PSA immunoassays.
Subject(s)
Antibodies, Monoclonal , Prostate-Specific Antigen/blood , Animals , Cross Reactions , Epitopes , Fluoroimmunoassay/methods , Humans , Kallikreins/analysis , Kallikreins/immunology , Male , Mice , Mice, Inbred BALB C , Prostate-Specific Antigen/immunology , Reagent Kits, Diagnostic , Tissue KallikreinsABSTRACT
We evaluated two chemical methods for quantifying mannitol in serum, based on the oxidation of mannitol by periodate, and measurement of the formaldehyde formed with chromotropic acid (colorimetry) or acetylacetone (fluorometry). We found interference in these methods by serum glycerol. Additionally, a high-performance liquid chromatography (HPLC) method was evaluated and found to be specific but impractical for routine use. We therefore, developed an enzymatic fluorometric procedure, based on the oxidation of mannitol by beta-NAD to fructose and NADH, in the presence of the enzyme mannitol dehydrogenase (MD). MD is not commercially available and was partially purified from cultures of Leuconostoc mesenteroides. This new method is specific, sensitive, simple, and accurate and is proposed as the method of choice for measuring mannitol in the serum of patients who received this sugar alcohol during routine hemodialysis treatment.
Subject(s)
Formaldehyde/analysis , Mannitol/blood , Renal Dialysis , Chromatography, High Pressure Liquid , Colorimetry , Evaluation Studies as Topic , Fructose/metabolism , Humans , Mannitol Dehydrogenases/metabolism , NAD/metabolism , Oxidation-Reduction , Pentanones/analysis , Periodic Acid/chemistry , Sensitivity and Specificity , Spectrometry, FluorescenceABSTRACT
Renal function was assessed in 42 stable outpatients who had been taking lithium for an average of 4 1/2 years. Impaired ability to concentrate the urine was found in 61% of the 41 patients who provided a urine sample for an osmolality measurement, and a moderate reduction in creatinine clearance was present in 12% of the entire group; 1 patient showed both defects. Urine microscopy revealed an excess of cells in 40%. It is suggested that lithium therapy produces a self-limiting lesion of the distal nephron that does not usually progress to chronic renal failure. The lesion is not dangerous, except that it may predispose to acute neurotoxic effects in the event of intercurrent illness or dehydration.
