Evaluation of a freeze-clamping technique designed for two- and three-dimensional metabolic studies of rat liver in vivo. Quenching efficiency and effect of clamping on tissue morphology

From identification of fluorescent flavoproteins to mitochondrial redox indicators in intact tissues

“…31 Since then, redox mapping of freeze-trapped tissues in two and three dimensions have been successfully applied to liver by exposing consecutive layers of frozen tissue for scanning. 32,33 The NADH/°avin°uorescence has gradually obtained more general use in biomedical research, and \normalized optical redox ratio" modi¯cations such as Flavin/(Flavin þ NADH) or NADH/(Flavin þ NADH) have been proposed to be less a®ected by hemodynamic changes or mitochondrial population density. 11,34 These modi¯ed ratios produce data which better obey the normal distribution 35 and also compress them to a zero-toone range.…”

Simple techniques for freeze clamping and for cutting and milling of frozen tissue at low temperature for the purpose of two- or three-dimensional metabolic studies in vivo

“…The present paper describes a number of techniques which have been developed as tools in such regional metabolic studies, aimed at a comparison between anatomical structure and spatial biochemical organization of various body organsin vivo (5,(8)(9)(10)(11)(12)(13)(14).…”

“…than 250 pm at a time will decrease the quality of the cut, i.e., macroscopically visible cutter marks on the surface may be seen. A tray underneath the cutter collects about 80% of the milling dust which may be used for metabolite analysis (see later) (12). The buzz saw (see Fig.…”

“…The milling and cutting instruments described in this paper were designed for cutting frozen tissue samples at very low temperatures for different types of threedimensional metabolic studies in viva (8)(9)(10)(11)(12)(13)(14). The techniques provide an alternative to microtome cutting which is usually used for such studies (2,5).…”

Manually Operated Freeze–Clamping Press

“…However, the DNA detection technologies, whether using enzymatic amplification of specific targets (4 -6) or hybridization and subsequent detection of a label (7), require complex enzymatic and processing steps, and this limits their use in field settings or where laboratory support is not available. Latex agglutination, widely used in immunoassay (8,9), has also been described for use in DNA hybridization assays (10,11). Here I describe a simple format that can detect hybridization using latex particle reagents alone, to low picogram sensitivities.…”