A rapid and simple method for the separation of pure lymphocytes from horse blood.
Abstract: A method for the separation of pure and viable lymphocytes and granulocytes from the same blood sample in horses was reported. By centrifuging equine heparinized blood at 100 xg for 10 min at room temperature (r.t.), the resulting supernatant plasma was an almost pure (97.71 +/- 0.30%; n = 15) suspension of highly viable (98.72 +/- 0.28%) lymphocytes. When sodium citrate was used as an anticoagulant, lymphocyte suspensions collected in the same manner showed lower purity (87.89 +/- 1.59%; n = 9) and higher yields (56.56 +/- 3.89%, n = 9 versus 36.11 +/- 2.23%, n = 15). Where needed, a further centrifugation at 250 xg for 3 min (r.t.) of heparinized lymphocyte preparations removed an average of 87.39% (n = 15) contaminating platelets. A suspension of 85.96 +/- 2.20% pure granulocytes (93.23 +/- 1.74% neutrophils; n = 14) with minimal contamination by erythrocytes and high viability (93.11 +/- 1.26%) was obtained by performing a flash red blood cell lysis on the white-greyish layer resulting from the centrifugation of the heparinized blood samples. Among the several methods available, the procedure described herein is easy, rapid, cheap and reproducible.
Publication Date: 2002-09-05 PubMed ID: 12208055DOI: 10.1016/s0165-2427(02)00185-xGoogle Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research article presents a fast and simple method for extracting pure lymphocytes and granulocytes from horse blood, using a centrifuge process.
Research Methodology
- The researchers carried out their study by using equine heparinized blood and subjecting it to centrifugation at a force of 100 xg for a duration of 10 minutes, at room temperature. This centrifugation process resulted in a supernatant plasma which was found to consist almost entirely of lymphocytes, with a sensed purity of 97.71% and viability of 98.72%.
- The anticoagulant used in the experiment also influenced the results. When sodium citrate was employed as an anticoagulant instead of heparin, the lymphocyte suspensions obtained from the centrifugation process were found to be of lower purity (87.89%) but of higher yields (56.56%) when compared to the use of heparin (36.11%).
- For the removal of contaminating platelets, researchers subjected heparinized lymphocyte preparations to further centrifugation at a force of 250 xg for 3 minutes. This was found to eliminate, on average, 87.39% of the contaminating platelets.
Additional Findings
- The study was able to also derive a suspension made up of 85.96% pure granulocytes with minimal contamination by red blood cells. This was obtained by performing a quick red blood cell lysis on the centrifuged heparinized blood samples. The vast majority of these granulocytes were neutrophils (93.23%), and had a high viability (93.11%).
- Significantly, the researchers concluded that among the several methods available for separating lymphocytes from blood, the procedure they used in this study is highly convenient: it is easy to use, quick, cost-effective, and reproducible.
Importance and Applications of the Study
- This study and its findings are of significant importance, particularly for research or treatment procedures that require the extraction of pure and viable lymphocytes and granulocytes from the blood. Such procedures could be applied across a range of scientific and medical fields including immunology, haematology and veterinary medicine.
- Moreover, the simplicity, speed, and affordability of this method make it accessible and manageable for wider applications, thus contributing to the advancement of these related fields.
Cite This Article
APA
Zizzadoro C, Belloli C, Badino P, Ormas P.
(2002).
A rapid and simple method for the separation of pure lymphocytes from horse blood.
Vet Immunol Immunopathol, 89(1-2), 99-104.
https://doi.org/10.1016/s0165-2427(02)00185-x Publication
Researcher Affiliations
- Department of Animal Health and Welfare, Division of Veterinary Pharmacology and Toxicology, University of Bari, Strada Provinciale per Casamassima Km 3, I-70010 Bari, Italy.
MeSH Terms
- Animals
- Blood Cell Count / veterinary
- Centrifugation / methods
- Centrifugation / veterinary
- Female
- Granulocytes / cytology
- Heparin / immunology
- Horses / blood
- Lymphocytes / cytology
- Male
Citations
This article has been cited 1 times.- Futas J, Horin P. Natural killer cell receptor genes in the family Equidae: not only Ly49. PLoS One 2013;8(5):e64736.
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