The construction of a microscope perfusion respirometer (MPR) for simultaneous recording of cellular respiration and microscopic morphology is described. All light microscope techniques for living cells (e.g. phase contrast, differential interference contrast (DIC), fluorimetry) can be applied to the monolayer cells grown on a coverslip. The main constituents of the MPR are a) a precision operating perfusion pump (constant volume output), b) a modified Dvorak-Stotler perfusion chamber, c) a special holder for the Clark-type oxygen probe, d) gas-tight connections of stainless steel tubing with dead volume-free fittings, and e) a temperature control unit. The cell material, established XTH (Xenopus laevis tadpole heart) cell is characterized. Examples of operation are presented, concerning a) normal respiration, b) respiration during uncoupling of oxidative phosphorylation by carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), and c) lactate production under anoxia. The corresponding mitochondrial in situ-morphology is demonstrated on photomicrographs. Details of construction and application are discussed. This new technique is supposed to extend the use of cell cultures instead of animal experiments in pharmaceutical routine tests.
In modern ultramicrotomes the thermic advance system has been replaced by a mechanical one. Earlier, conventional DC-voltage motors were used, but recently the step motor has become popular. In connection with such motors, the application of digital control elements is facilitated, thus increasing precision significantly. A further step towards improving the precision of the advance system is the application of microprocessors as intelligent and interacting control elements. The microprocessor can not only take over the function sensors, the function of regulation. This paper describes a working concept whereby the advance system does not, as usual, work freely, but in connection with a sensor which measures the advance, and first after a positive data comparison over the microprocessor, allows the cutting movement. The concept is not limited to ultramicrotomes but can also be applied to mechanical rotation microtomes.
This paper presents a method for labeling antibodies with fluorescein isothiocyanate (FITC) and additional with horseradish peroxidase (HPOD). With this double labelled antisera the fluorescence-serologic antibody technique (FAT) as well as the enzyme-serologic antibody technique (EAT) was done bit by bit for the same object. How the presented pictures demonstrate the result of both techniques, FAT and EAT, are the same with the described method. So, it is possible after fluorescence-microscopical exploitation through a following histochemical proof of the HPOD to get permanent preparations with the same biological result.
A simple apparatus for the processing of biological specimens for electron microscopy is described. It comprises a fluid exchange chamber with associated reservoir. By carrying out all specimen processing from fixation to embedding in the exchange chamber and infiltrating the dehydrating and embedding mediums progressively, the handling and osmotic stresses associated with standard step processing are minimised.
