Improved enrichment of insect gap junctions by filtration and sonication of NaOH-extracted subcellular fractions

The purification of gap junctions from insects has been hampered by low yields when starting with dissected tissues or by contamination with non-junctional structures when starting with intact insects. A method is described here involving filtration and sonication of NaOH-extracted crude membrane fractions from larvae of the lepidopteran Heliothis virescens which yields fractions containing approximately 50% gap junctions and approximately 7 microg total protein per g of larvae and represents an estimated 10-100 fold increase in gap junction enrichment compared with a previously described procedure (Cell Tissue Res. 274: 393-403, 1993). The remaining structures in the fractions consist of non-junctional membrane, septate junctions, lamellate bodies and amorphous material.