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Neutrophil isolation from human peripheral blood

DA Dhia Azzouz
MK Meraj A. Khan
NP Nades Palaniyar
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Peripheral blood drawn from healthy donors was placed in K2 EDTA blood collection tubes. Neutrophils were isolated as previously published using PolymorphPrep4. Modifications to the manufacturer’s neutrophils isolation protocol were made. Red blood cell lysis step was added by using a hypotonic solution of 0.2% (w/v) NaCl. The solution was then made isotonic and buffered by adding an equal volume of 1.6% (w/v) NaCl solution with Hepes buffer (20 mM, pH 7.2). The cells were washed twice with a solution of 0.85% (w/v) NaCl and Hepes (10 mM, pH 7.2). Neutrophils were then resuspended in RPMI medium (Invitrogen) containing Hepes buffer (10 mM, pH 7.2).

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