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Elimination of paraprotein interference in determination of plasma inorganic phosphate by ammonium molybdate method
Z Zaman1, L Sneyers, A Van Orshoven
1Department of Clinical Chemistry, University Hospitals Leuven, Belgium.
Abstract:
Phosphate concentrations were determined in 52 cases of paraproteinemia. The unmodified acidic ammonium molybdate method produced 19% spuriously high results. The false increase of phosphate concentration was attributable to formation of precipitate in the reaction mixture. The precipitate was formed by interaction between immunoglobulins and the unmodified acidic ammonium molybdate reagent. The magnitude of interference bore no relation to the type, concentrations, or isoelectric point of the paraproteins or to the presence or absence of free light chains. Diluting the sample to approximately 40 g/L total protein reduced but did not always eliminate the interference. In some cases paraprotein concentration as low as 8 g/L falsely increased plasma phosphate results. Apparently, only IgG and IgM but not IgA paraproteins produced the interference. Deproteination by ultrafiltration or by treatment with trichloroacetic acid removed the interference. The Kodak slide method and the new modified Boehringer Mannheim phosphate test were found to be interference-free. However, in some cases the latter new formulation is sensitive to substantial changes in ionic concentration of the reaction mixture.
Insights
The unmodified acidic ammonium molybdate method for phosphate determination can yield falsely high results in paraproteinemia due to immunoglobulin interference. Alternative methods like the Kodak slide test or modified Boehringer Mannheim assay are recommended for accurate phosphate measurement.
Area of Science:
- Clinical Chemistry
- Immunology
- Biochemistry
Background:
- Paraproteinemia involves abnormal immunoglobulin production, potentially affecting laboratory test results.
- Accurate phosphate measurement is crucial for diagnosing and managing various metabolic and renal conditions.
Purpose of the Study:
- To investigate the interference of paraproteins in plasma phosphate concentration determination using the unmodified acidic ammonium molybdate method.
- To identify the characteristics of paraproteins causing interference and evaluate methods to mitigate or avoid it.
Main Methods:
- Phosphate concentrations were measured in 52 paraproteinemia cases using the unmodified acidic ammonium molybdate method.
- Interference was assessed based on paraprotein type, concentration, isoelectric point, and presence of free light chains.
- Sample dilution, deproteination (ultrafiltration, trichloroacetic acid), and alternative assay methods (Kodak slide, modified Boehringer Mannheim) were evaluated.
Main Results:
- The unmodified acidic ammonium molybdate method yielded spuriously high phosphate results in 19% of cases.
- Precipitate formation due to immunoglobulin-reagent interaction caused the interference, independent of paraprotein characteristics.
- Interference was observed with IgG and IgM paraproteins, but not IgA.
- Deproteination and specific alternative methods (Kodak, modified Boehringer Mannheim) eliminated interference, though the latter showed sensitivity to ionic concentration.
Conclusions:
- The unmodified acidic ammonium molybdate method is unreliable for phosphate determination in paraproteinemia.
- Accurate phosphate measurement in these patients requires methods free from immunoglobulin interference.
- Modified Boehringer Mannheim and Kodak slide methods are viable alternatives, with consideration for ionic concentration effects in the former.