ISOLATION AND IMMOBILIZATION OF VARIOUS PLASTID SUBTYPES BY MAGNETIC IMMUNOABSORPTION

Antibodies have been prepared which immunolocalize to the outer membra ne of the pea chloroplast envelope and cause agglutination of isolated chloroplasts. This antisera is immunoreactive with a variety of plast id forms from both monocotyledonous and dicotyledonous plants. Whether such antibodies might be effectively used for isolation and immobiliz ation of plastids from whole cell lysates has been tested. A system ha s been developed for immunolabeling various forms of higher plant plas tids with biotinylated antibody and streptavidin magnetic nano-particl es followed by separation of the plastids in a 0.6 Tesla high gradient magnetic field. Using this magnetic immunoabsorption procedure it has been possible to achieve a high degree of positive enrichment for chr omoplasts, amyloplasts, and chloroplasts from whole cell lysates of se veral plant species. The integrity of these plastids has been examined by in organellar protein synthesis, C-14-ADP-glucose uptake, flow cyt ometry, in vitro synthesized precursor import and FITC-cationized ferr itin staining of the plastid envelope. Western blot analysis showed si gnificant enrichment for amyloplasts from cytosolic sucrose synthase i n maize endosperm. Magnetic immunoabsorption of subcellular structures from whole cell lysates is a new method that may be useful in the in vitro analysis of many different cellular compartments from a wide ran ge of organisms.