Subcellular fractionation of human neutrophils on Percoll density gradients

L Kjeldsen, H Sengelov, N Borregaard

Abstract

Subcellular fractionation has been an important tool in the investigation of neutrophil structural organization including granule heterogeneity, composition and mobilization. The resolution of organelles obtained by subcellular fractionation was improved considerably after the introduction of nitrogen cavitation as an efficient but gentle means of disrupting neutrophils and with Percoll as a density medium. This paper describes in detail the methodology of subcellular fractionation of nitrogen cavitated neutrophils on one-, two-, and three-layer Percoll density gradients. Appropriate marker proteins are presented for neutrophil organelles including azurophil, specific and gelatinase granules, in addition to secretory vesicles and plasma membranes. The dynamics of granule and secretory vesicle exocytosis is demonstrated by subcellular fractionation of resting and activated human neutrophils. Finally, the paper describes the applications of subcellular fractionation in the investigation of the localization of neutrophil constituents, in protein purification schemes and in the study of translocation of cytosolic proteins to isolated neutrophil organelles.

OriginalsprogEngelsk
TidsskriftJournal of Immunological Methods
Vol/bind232
Udgave nummer1-2
Sider (fra-til)131-43
Antal sider13
ISSN0022-1759
DOI
StatusUdgivet - 17 dec. 1999
Udgivet eksterntJa

Fingeraftryk

Dyk ned i forskningsemnerne om 'Subcellular fractionation of human neutrophils on Percoll density gradients'. Sammen danner de et unikt fingeraftryk.

Citationsformater