Dissociation of lepidopterous tissues with proteolytic enzymes: Comparison of collagenase,trypsin, and pronase

The fat body and certain other tissues of lepidopterous larvae and diapausing pupae can be completely dissociated by incubation in crude or purified collagenase. Damage to the cells is minimal. Dissociation with trypsin is incomplete, and pronase causes extensive damage. All three enzymes act principally by digesting the extracellular connective sheath that envelops the individual lobes of fat body, since the cells at this stage are not intrinsically cohesive. The experimental findings support the notion that collagen is an important structural component of insect connective membranes.     

Purification and characterization of chick intestine brush border membrane Effects of 1α(OH) vitamin D3 treatment

A new technique has been developed for the isolation of membrane vesicles from the vitamin D-deficient and vitamin D-treated chick intestinal brush border membrane. The technique involves removal of nuclei from a low speed pellet by discontinuous sucrose gradient centrifugation. The resulting intact brush borders are then homogenized in 0.5 M Tris and the membrane fragments purified on a glycerol gradient. This preparation represents a 20-fold purification of the brush border marker sucrase. After 1α-hydroxyvitamin D₃ treatment there is a significant increase in membrane phospholipid phosphorous, an alteration in the fatty acid composition of the phosphatidylcholine fraction of membrane phospholipid, and a decrease in sucrase specific activity.     

Xanthine oxidase,an indicator of secretory differentiation in mammary cells

Elevated levels of xanthine oxidase were found in (1) lactating mouse mammary glands, compared with virgin and midpregnant glands; and (2) primary mouse mammary cells cultured on floating collagen gels, compared with non-secretory cells on attached gels. In primary culture, increase in xanthine oxidase activity above a basal level coincided with secretory activity as measured by casein production; intracellular levels of casein and xanthine oxidase showed a high degree of correspondence. It is suggested that xanthine oxidase levels can be used as an indicator of in vivo and in vitro secretory differentiation in mammary epithelial cells.