Immunoaffinity purification and immunoassay determination of human erythrocyte acylphosphatase.
Specific anti-human erythrocyte acylphosphatase antibodies were raised in rabbits, purified by affinity chromatography, and used to develop an enzyme purification procedure based on an immunoaffinity chromatography step. This procedure permitted the rapid purification of the enzyme, with a high final yield and with a specific activity very similar to that found for the enzyme purified by the standard procedure. The noncompetitive enzyme-linked immunoadsorbent assay developed with the affinity-purified antibodies was very specific and sensitive in that a positive reaction could be detected in the presence of antigen amounts of as little as 0.01 ng/ml. By this assay the enzyme content was determined in normal cells, tissues, and organs as well as in blood samples from hemopathy-affected patients. This test could possibly have clinical applications.
Degl'Innocenti
D
D
Department of Biochemical Sciences, University of Florence, Italy.
Berti
A
A
Stefani
M
M
Liguri
G
G
Ramponi
G
G
D017766:Acid Anhydride Hydrolases; D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000302:isolation & purification; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007118:Immunoassay; D007527:Isoenzymes / Q000378:metabolism; D009928:Organ Specificity; D010744:Phosphoric Monoester Hydrolases / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D011817:Rabbits; D015203:Reproducibility of Results; D000097024:Acylphosphatase
D017766:Acid Anhydride Hydrolases; D000906:Antibodies; D007527:Isoenzymes; D010744:Phosphoric Monoester Hydrolases; D000097024:Acylphosphatase