Production and certification of an enzyme reference material for pancreatic α-amylase (CRM 476)

Gemma Gubern, Francesca Canalias, F. Javier Gella, Elizabeth Colinet, Christos Profilis, Derek H. Calam, Ferruccio Ceriotti, J. Dufaux, Anthony G. Hadjivassiliou, Jean Marc Lessinger, Klaus Lorentz, Anne Vassault

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

We describe the preparation of a lyophilized material containing purified human pancreatic α-amylase and the certification of its catalytic concentration. The enzyme was purified from human pancreas by ammonium sulphate precipitation and chromatography successively on DEAE-Sephacel, CM-Sepharose and Sephadex G-75. The purified enzyme had a specific activity of 52.9 kU/g protein and was 299% pure on polyacrylamidegel electrophoresis. Only trace amounts of lipase and lactate dehydrogenase were detected in the purified fraction. The purified pancreatic α-amylase had a molar mass of 57 500 g/mol and an isoelectric point at 7.1. The material was prepared by diluting the purified α-amylase in a matrix containing PIPES buffer 25 mmol/l, pH 7.0, sodium chloride 50 mmol/l, calcium chloride 1.5 mmol/l, EDTA 0.5 mmol/l and human serum albumin 30 g/l, dispensing in ampoules and freeze-drying. The ampoules were homogeneous and the yearly loss of activity on the basis of accelerated degradation studies was less than 0.01% at -20°C. The certified value for α-amylase catalytic concentration in the reconstituted reference material is 555 U/l ± 11 U/l when measured by the specified method at 37°C. The material can be used to verify the comparability of results from different laboratories, for intra-laboratory quality control or for calibration of α-amylase catalytic concentration measurements.

Original languageEnglish
Pages (from-to)145-162
Number of pages18
JournalClinica Chimica Acta
Volume251
Issue number2
DOIs
Publication statusPublished - Jul 30 1996

Fingerprint

Certification
Amylases
Enzymes
Calcium Chloride
Freeze Drying
Molar mass
Isoelectric Point
Ammonium Sulfate
Chromatography
Electrophoresis
Lipase
L-Lactate Dehydrogenase
Sodium Chloride
Serum Albumin
Edetic Acid
Quality Control
Sepharose
Calibration
Quality control
Pancreas

Keywords

  • Enzyme activity
  • Refeence material
  • Standardization

ASJC Scopus subject areas

  • Biochemistry
  • Clinical Biochemistry

Cite this

Gubern, G., Canalias, F., Gella, F. J., Colinet, E., Profilis, C., Calam, D. H., ... Vassault, A. (1996). Production and certification of an enzyme reference material for pancreatic α-amylase (CRM 476). Clinica Chimica Acta, 251(2), 145-162. https://doi.org/10.1016/0009-8981(96)06302-4

Production and certification of an enzyme reference material for pancreatic α-amylase (CRM 476). / Gubern, Gemma; Canalias, Francesca; Gella, F. Javier; Colinet, Elizabeth; Profilis, Christos; Calam, Derek H.; Ceriotti, Ferruccio; Dufaux, J.; Hadjivassiliou, Anthony G.; Lessinger, Jean Marc; Lorentz, Klaus; Vassault, Anne.

In: Clinica Chimica Acta, Vol. 251, No. 2, 30.07.1996, p. 145-162.

Research output: Contribution to journalArticle

Gubern, G, Canalias, F, Gella, FJ, Colinet, E, Profilis, C, Calam, DH, Ceriotti, F, Dufaux, J, Hadjivassiliou, AG, Lessinger, JM, Lorentz, K & Vassault, A 1996, 'Production and certification of an enzyme reference material for pancreatic α-amylase (CRM 476)', Clinica Chimica Acta, vol. 251, no. 2, pp. 145-162. https://doi.org/10.1016/0009-8981(96)06302-4
Gubern, Gemma ; Canalias, Francesca ; Gella, F. Javier ; Colinet, Elizabeth ; Profilis, Christos ; Calam, Derek H. ; Ceriotti, Ferruccio ; Dufaux, J. ; Hadjivassiliou, Anthony G. ; Lessinger, Jean Marc ; Lorentz, Klaus ; Vassault, Anne. / Production and certification of an enzyme reference material for pancreatic α-amylase (CRM 476). In: Clinica Chimica Acta. 1996 ; Vol. 251, No. 2. pp. 145-162.
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