Lipids tightly bound to cell and tissue proteins (lipoproteins) were investigated with the acetone-Sudan procedure in cryostat-cut sections of normal and regenerating limb tissues of the adult newt, Diemictylus viridescens.Nuclear and cytolplasmic membranes of all tissues stained moderately black; the nucleo-and cytoplasm stained less intensely than their membranes. Connective tissue fibers of the dermis, mysial and neural sheaths, and tunics of blood vessels yielded intense responses, In striated muscle of the limb, myofibrils and associated striations responded strongly. Nerve myelin responded weakly.In the preblastemic regenerate, the fibrous adepidermal basement membrane terminated abruptly at the surface of amputation. A distal fibrocellular residue was evident in continuity with the retrograde degeneration of amputation-injured muscle: observations suggest a possible contribution to the fibrocellular reticulum from myofibrils. The regeneration blastema appears isolated from proximal limb tissues by the intervening fibrocellular reticulum. The response of the blastema cell is relatively weaker than that of other cells and tissues. Regenerating muscle is recognized by the appearance of prominently stained myofibrils in myoblastic extensions off the limb stump musculature. The matrix and chondrocytes of regenerating cartilage stain for lipoprotein, with the osteoid of osteogenic centers responding strongly.Polychrome responses were obtained from hyalin-bodies within interphase nuclei, and from chromosomes in mitosis, suggesting that chromosomal lipid has been stained.Supporting the lipidic character of the observed responses is the negative reaction following long term lipid extraction in warm chloroform: methanol.Histochemical studies are currently being conducted on the lipids of the normal and regenerating tissues of the adult newt forelimb, based on the suggestion by Schmidt and Weidman ('64) that lipids may play a significant role in the events of regeneration. Previous studies have revealed a prominent sudanophilic lipid response (Schmidt, '66a), acidic and nonacidic lipid, phosphatides, and stainable cholesterol (Schmidt, '66b) in these tissues. However, the characterization of the tissue lipids, especially that of the regeneration blastema, remains incomplete. Therefore, we undertook the study of socalled masked lipids, or lipoproteins, in the normal and regenerating tissues of the adult newt limb: preliminary studies have been reported (Schmidt, '64).Lipids that are tightly bound to tissue proteins have been demonstrated by various histochemical procedures, e.g., Ackerman, '52; Berenbaum, '58; Ciacco, '34; Serra, '58; and others. These methods are founded on the prior or simultaneous application of some unmasking agent. Of the several possible procedures we employed, the lipoprotein responses which are reported herein are based exclusively on the acetone-Sudan test of Berenbaum ('58). Warm acetone serves as the unmasking agent and sudan black B as the lipid indicator. Although objections have ...
