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A two-step immunomagnetic separation of somatic cell subpopulations for a gene expression profile study in bovine milk

Published online by Cambridge University Press:  08 August 2018

Sara Divari ,
Laura Starvaggi Cucuzza ,
Fulvio Riondato ,
Paola Pregel ,
Paola Sacchi ,
Roberto Rasero ,
Bartolomeo Biolatti  and
Francesca Tiziana Cannizzo
Sara Divari*
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Laura Starvaggi Cucuzza
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Fulvio Riondato
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Paola Pregel
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Paola Sacchi
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Roberto Rasero
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Bartolomeo Biolatti
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
Francesca Tiziana Cannizzo
Affiliation:
Department of Veterinary Science, University of Turin, Largo Braccini 2, 10095 Grugliasco, Turin, Italy
*
*For correspondence; e-mail: [email protected]
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Abstract

The objective of this study was to demonstrate the usefulness of an immunomagnetic method to purify subpopulations of milk somatic cells. The experiment was conducted on milk samples collected from healthy cows (n = 17) and from cows with clinical mastitis (n = 24) due to a Staphylococcus aureus natural infection. A two-step immunomagnetic purification was applied to simultaneously separate three somatic cell subpopulations from the same milk sample. Total RNA was extracted and qPCR was performed to determinate mRNA levels of innate immunity target genes in purified somatic cell subpopulations. Good quality and quantity of RNA allowed the reference gene analysis in each cell subpopulation. An up-regulation of the main genes involved in innate immune defence was detected in separated polymorphonuclear neutrophilic leucocytes-monocytes and lymphocytes of mastitic milk. These results and flow cytometric analysis suggest that the immunomagnetic purification is an efficient method for the isolation of the three populations from milk, allowing the cells to be studied separately.

Keywords

Mastitisinnate immunitygene expressionmilk somatic cellsmagnetic separation
Type
Research Article
Information
Journal of Dairy Research , Volume 85 , Issue 3 , August 2018 , pp. 281 - 287
Copyright
Copyright © Hannah Dairy Research Foundation 2018 

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