Impact of pronase on flow cytometric crossmatch outcome

Sarah J. Hetrick, Karl P. Schillinger, Andrea A. Zachary, Annette Jackson

Research output: Contribution to journalArticle

Abstract

Pronase treatment of lymphocytes is used to reduce nonspecific binding of immunoglobulins in flow cytometric crossmatch (FCXM) tests and at higher concentrations to remove CD20 from the cell surface. We examined the effect of pronase treatment on human leukocyte antigen (HLA) expression and on FCXM results. Lymphocytes were tested untreated and after treatment with either 2 mg/mL (10 cell donors) or 1 mg/mL (6 cell donors) of pronase. The 2 mg/mL concentration reduced HLA expression in 28 of 30 (93%) cases. The reduction was statistically significant for HLA class I antigens on T cells (33 - 10%, p = 0.0006), class I on B cells (23 - 13%, p = 0.012), and class II on B cells (45 - 37%, p = 0.005). FCXMs were performed using pronase-treated and untreated cells. The 2 mg/mL concentration of pronase reduced reactivity in 5 of 16 (31%) tests of T cells and 15 of 16 (94%) tests of B cells. Of the remaining 11 T-cell tests, the reactivity was unchanged (*10% difference) in 5 and increased by 18-73% in 6. Treatment with 1 mg/mL of pronase significantly increased reactivity in 20 of 23 tests of T cells (87%, p = 6.0 × 10-5). These data indicate that pronase treatment may result in erroneous FCXM results.

Original languageEnglish (US)
Pages (from-to)330-336
Number of pages7
JournalHuman Immunology
Volume72
Issue number4
DOIs
StatePublished - Apr 2011

Fingerprint

Pronase
HLA Antigens
T-Lymphocytes
B-Lymphocytes
Lymphocytes
Histocompatibility Antigens Class I
Immunoglobulins

Keywords

  • Flow cytometric crossmatch
  • HLA expression
  • Pronase

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

Hetrick, S. J., Schillinger, K. P., Zachary, A. A., & Jackson, A. (2011). Impact of pronase on flow cytometric crossmatch outcome. Human Immunology, 72(4), 330-336. https://doi.org/10.1016/j.humimm.2011.01.005

Impact of pronase on flow cytometric crossmatch outcome. / Hetrick, Sarah J.; Schillinger, Karl P.; Zachary, Andrea A.; Jackson, Annette.

In: Human Immunology, Vol. 72, No. 4, 04.2011, p. 330-336.

Research output: Contribution to journalArticle

Hetrick, SJ, Schillinger, KP, Zachary, AA & Jackson, A 2011, 'Impact of pronase on flow cytometric crossmatch outcome', Human Immunology, vol. 72, no. 4, pp. 330-336. https://doi.org/10.1016/j.humimm.2011.01.005
Hetrick SJ, Schillinger KP, Zachary AA, Jackson A. Impact of pronase on flow cytometric crossmatch outcome. Human Immunology. 2011 Apr;72(4):330-336. https://doi.org/10.1016/j.humimm.2011.01.005
Hetrick, Sarah J. ; Schillinger, Karl P. ; Zachary, Andrea A. ; Jackson, Annette. / Impact of pronase on flow cytometric crossmatch outcome. In: Human Immunology. 2011 ; Vol. 72, No. 4. pp. 330-336.
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