Natural killer cells(NK cells) are currently under intensive investigation because of their possible role in immune survilunce against malignancy, resistance against microbial infections and regulation of lymphohematopoiesis.
Several methods such as sheep erythrocyte rosette method, passing the nylon-wool or IgG-anti IgG column after removal of adhesive macrophage or phagocytic monocytes for isolation and measurement of natural cytotoxicity of peripheral plood lymphocytes have been used.
Recently human NK cells as a morphological subpopulation of white blood cells, called large granular lymphocytes (LGL) by a relative high cytoplasmic: nuclear ratio, azurophilic cytoplasmic granules and an often indented eccentric nucleus, have been identified by discontinous percoll density gradient centrifugation method which is relatively simple and less time spending method.
In order to find out the possibility of seperation of LGL-riched lymphocytes by discontinous percoll densitygradient centrifugation and to measure natural cytotoxicity from normal peripheral blood, authors observed the osmolarity of percoll solution, recovery rates of lymphocyte and distribution of LGL after removal of adhesive macrophages, phagocytic monocytes and percoll density gradient centrifugation with its´ natural cytotoxicity.
The obtained results were as follows;
1) The osmolarities of 37.5% to 50% percoll solutions(10X PBS+RPMI-1640-FBS+Percoll) were 289.4±1.02~302.0±63 mOmol/kg H₂O.
2) The distribution of LGL after removal of adhesive macrophage, phagccytic monocytes were 8.6±1.85%, 11.2±2.64%. But LGL of fraction 1,2 and 3(47.2±6.40%, 65.2±4.99% and 36.8±4.83%) were significantly increaesd after percoll density gradient centrifugation.
3) The natural cytotoxicity of LGL-riched lymphocytes of fraction 1, 2 and 3(65. 1±10. 73%, 74.5±3.89% and 54.9±8.19%) by percoll density gradien centrifugation were significantly higher than that of removing adhesive macrophaLe(8.9±2.38%) or phagocytic monocytes(24.5±11.41%). But fraction 4 to 7(25.5±3.17%~5.7±1.17%) revealed low cytotoxicity.
With above results, it isinferred that LGL-riched lymphocytes, which having relatively high natural cytotoxicity, can be isolated by discontinous percoll density gradient centrifugation which is relatively simple, less time spending method.
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