Cacodylate buffer, solution preparation

Two-thirds of the blocks of 1 mm diameter were immersed for 12 h in a ZIO mixture prepared according to the method of Maillet (1), and was first utilized in electron microscopy by Stach (15a). The remaining blocks were prefixed at 4°C for 2 h in 0.1 M cacodylate buffer containing 5% glutaraldehyde for 15 min, and immersed in the ZIO mixture at 25°C for 12 h. The ZIO mixture was prepared as follows twelve to 15 g of zinc (powder) and 5 g of iodine (crystal) were dissolved in 200 mL of distilled water. Eight milliliters of the filtered solution was added to 2 mL of 2% Os04 solution prior to use. 

The tissues were fixed in 0.05 M cacodylate buffer containing 2.5% glutaraldehyde and 1.5% formaldehyde (pH 7.0) for 16 h. The ZIO mixture was prepared as follows 3 g zinc (powder) and 1 g resublimed iodine crystals were dissolved in 20 mL distilled water. After stirring for 5 min, the zinc was filtered off. The filtered solution was mixed with an equal volume of 2% 0s04 solution and the solution used immediately. Treatment with the ZIO mixture was carried out for 4 h at room temperature. 

After primary fixation the algal and root samples were rinsed in 0.1 M PIPES buffer, pH 7.0, then twice in 0.1 M sodium cacodylate buffer, pH 6.8, and post-fixed in 1% osmium tetroxide in 0.1 M cacodylate buffer, pH 6.8, overnight at 4°C. The osmium solution prepared for the root samples was added with 0.7% potassium ferricyanide in order to improve osmium penetration in the root tissues. This was particularly necessary for the 2 h control roots and roots from RO-treated seed, due to their relatively low water content. The replacement of PIPES buffer with cacodylate buffer before osmication was necessary, as PIPES reacts with osmium, producing a dark precipitate. 

Cacodylic acid, (CH3)2As02H, is a toxic compound that behaves as a weak acid. It is used to prepare buffered solutions. For the following reaction pKa = 6.19 ... 

Buffer system according to Davies and Rosenbaum [178]. This buffer system has been worked out for preparative purposes. 100 ml of acrylamide solution (prepared from 3.67% acrylamide, 5% bisacrylamide, in 0.012 M Tris, 0.013 M cacodylate (pH... 

When all the ovaries have been dissected, transfer them to a 100- il drop of fixative (5% formaldehyde made by diluting freshly prepared 37% solution in cacodylate buffer, prewarmed to 37"C) on the dissecting surface. Start a timer set for 4 minutes. 

In a prior publication, initial measurements of ket were reported for tris/cacodylate buffers of pH 6-8.5 and ionic strengths of 1-100 mM. The cytochrome c samples in those experiments had been chromatographically purified according to established procedures and then lyophilized for subsequent storage at -18 C. Solutions were then prepared directly from the lyophilized material. Since lyophilization has been shown to have very deleterious effects on the silver electrochemistry of cytochrome c and also results in the appearance of new chromatographic bands, we have repeated our earlier experiments using purified, non-lyophilized samples. 

Fixative solution Sodium cacodylate buffer with SMB, 0.65 % glutaraldehyde see Note 2). Prepare the amount needed always freshly. 

Dilute the RNA solution (10—50 jxM) in CEK buffer (10 mM potassium cacodylate, 0.1 mM EDTA at pH 7.3, 100 mM KC1) for Mg2-1-titration or in 1 X CE buffer (for monovalent ion titration). We consis-tendy get good results with RNA concentrations corresponding to an absorbance of 0.5—0.6 O.D. Prepare RNA in appropriate amounts for up to three folding data points for each centrifuge run. 

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