title: Determination of N-acetyl-β-hexosaminidase in serum from hemolyzed blood.
abstract: BACKGROUND
  Determination of lysosomal N-acetyl-β-hexosaminidase (HEX) in serum from hemolyzed blood, creates serious analytical problems, because hemoglobin absorbs light at a similar wavelength like 4-nitrophenol, which is released from artificial substrate.
  
  
  OBJECTIVE
  The objective of the work was to adapt a manual method to allow analysis of HEX in hemolyzed samples.
  
  
  METHODS
  Serums without and with hemolysis were incubated with 4-nitrophenol-N-acetylglucosamine as a substrate. Released 4-nitrophenol was determined colorimetrically. After the incubation of the serum from hemolyzed blood with substrate, hemoglobin was precipitated with trichloroacetic acid (TCA) before 4-nitrophenol determination.
  
  
  RESULTS
  The mean concentration of HEX activity in non-hemolyzed and hemolyzed blood of the same patients, determined with non-modified and modified methods had no significant differences, and they are: 243.12±119.76 and 233.99±108.76pkat/mL, respectively. A coefficient of correlation between non-modified and modified methods equals the 0.98. For HEX determination with the modified method in serum from hemolyzed blood, optimal reaction time was 60min, pH of reaction mixture was 4.7, and Km was 0.11mMm.
  
  
  CONCLUSIONS
  HEX determinations in the same serums from non-hemolyzed blood by the non-modified method and hemolyzed blood with the modified method, gave similar results with a 0.98 coefficient of correlation. The modified method is appropriate for HEX determination in serum from hemolyzed blood.
authors.affiliation: Medical Institute, The College of Computer Science and Business Administration, Łomża, Poland. Electronic address: schojnowska@pwsip.edu.pl.
authors.forename: Sylwia
authors.initials: S
authors.lastname: Chojnowska
authors.affiliation: Department of Biochemistry, Radioimmunology and Experimental Medicine, The Children Memorial Health Institute, Warsaw, Poland.
authors.forename: Alina
authors.initials: A
authors.lastname: Kępka
authors.affiliation: Department of Emergency Medicine and Disasters, Medical University of Bialystok, Poland.
authors.forename: Sławomir Dariusz
authors.initials: SD
authors.lastname: Szajda
authors.affiliation: Łomża Medical College of the Universal Educational Society, Łomża, Poland.
authors.forename: Zbigniew Paweł
authors.initials: ZP
authors.lastname: Kołodziejczyk
authors.affiliation: Łomża Medical College of the Universal Educational Society, Łomża, Poland.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Zwierz
authors.affiliation: Department of Psychiatry, Medical University of Bialystok, Poland.
authors.forename: Napoleon
authors.initials: N
authors.lastname: Waszkiewicz
mesh_terms: D000293:Adolescent; D000328:Adult; D015415:Biomarkers / Q000097:blood*; D005260:Female; D005500:Follow-Up Studies; D006454:Hemoglobins / Q000032:analysis*; D006461:Hemolysis / Q000502:physiology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D055815:Young Adult; D001619:beta-N-Acetylhexosaminidases / Q000097:blood*
chemical_list: D015415:Biomarkers; D006454:Hemoglobins; D001619:beta-N-Acetylhexosaminidases
keywords: Hemolysis; N-acetyl-β-hexosaminidase (HEX); Serum