Veronal-acetate buffer

Veronal-acetate buffer pH 9.2 Sodiumacetate 30 mM, sodium barbitone or barbitone acid 30 mM, NaCl 100 mM, and MgCl2 50 mM. (Can be made five times the foregoing concentration.)... 

Developer Naphthol AS-BI phosphate (Sigma N4875) 2 mg, N,N-dimeth-ylformamid 0.2 mL, veronal-acetate buffer 9.8 mL, pH 9.2, levamisol 1 M... 

Wash in veronal-acetate buffer, pH 9.2, for 5 min. Dry with paper towels. 

Fix pollen tubes 30 min in 6.25% glutaraldehyde in phosphate buffer or veronal-acetate buffer, pH 7.4. 

Develop the alkaline phosphatase with fast red dissolve 5 mg sodium naphthol AS BI phosphate in dimethylformamide (few drops) and add to 5 mg fast red TR salt in 10 ml veronal acetate buffer (pH = 9.2), incubate the slides for 1 h Wash with tap water Counterstain as desired... 

Veronal acetate buffer 0.97 g of sodium acetate (trihydrate), 1.47 g of sodium barbitone, 250 mL of fresh distilled water (C02 free), and 2 5 mL of 0 M hydrochloric acid, pH 9.2... 

Fast red salt Dissolve 5 mg of naphthol AS BI phosphoric acid sodium salt m 1 drop of DMF in a glass tube. Dissolve 5 mg of Fast red TR salt in 10 mL of veronal acetate buffer, pH 9 2 Mix the two solutions together and filter... 

Phosphate-citrate buffer.b Phosphate-acetate buffer. High or low mouse strains (see Section II). Phthalate buffer. Veronal-acetate buffer. 

Counterstain if desired DAB lightly with hematoxylin or 0.1 % methyl green (chloroform washed) in veronal-acetate buffer, pH 4.0 HY with neutral red or carma-lum AEC with hematoxylin (lightly) since product is alcohol-soluble and differentiation of the hematoxylin with acid-alcohol should be avoided. 

Buffers. Best results, as a rule, are obtained at a pH on the alkaline side of the isoelectric points of the proteins in the mixture. The commonly used buffers include (i) 0.05 M veronal buffer at pH 8.6, and (ii) veronal/acetate buffer at pH 8.6 and ionic strength 0.1. 

Fig. 14. Electrophoretic homogeneity of porcine lipase (16). Starch columns (3.0 X 90 cm) equilibrated with 0.025 M acetate buffer, pH 5.25 (diagram on the left), or 0.025 Af veronal buffer, pH 8.0 (diagram on the right). Potential gradient inside the column, 8 volts/cm temperature, +1°C. Elution after 48 hr. Ordinates, per cent in 1 ml eluate of the total lipase activity (solid line) and total proteins (dotted line) introduced into the column. The vertical dotted line gives the true origin, with due regard to the electroosmotic flow. The figures along the peaks give the specific activity of some fractions. Specific activity of the sample introduced into the column, 3500. Abscissas, volume of eluate in milliliters.

Yamakawa (Yl) studied 150 juices by free boundary and paper electrophoresis in veronal buflFer of pH 8.6, phosphate buffer of pH 7.6, borate buffer of pH 5.8, and acetate buffer of pH 4.5. The normal electrophoretic pattern consisted of four to seven peaks, of which four were considered to be main components. The best resolution was obtained with the first three buffers. In gastric ulcer, the slowest component, B4, was increased whereas, in atrophic gastritis and gastric cancer, components B3 and B4 were high and additional subcomponents were noted. 

Wada et al. (W7) studied mucoprotein and mucoproteose fractions by horizontal paper electrophoresis in veronal buffer of pH 8.6 and in acetate buffer of pH 4.5. They processed these materials from gastric juices of normals, patients with histamine-fast anacidity, and those with gastric cancer, which were collected after insulin stimulation. They also subjected the trichloroacetic acid precipitate of gastric juice to electrophoresis, as well as the supernatant fraction remaining after acetone precipitation of the trichloroacetic filtrate of the gastric juice. 

L. Michaelis has reported also a combination of veronal and acetate buffers which, due to the addition of an appropriate amount of sodium chloride, have the same ionic strength as a salt solution isotonic with blood. The original solution is 1/7 molar with respect to sodium acetate and the sodium salt of veronal, 500 c.c. of solution (in carbon dioxide-free water) containing 9.714 g.-of sodium acetate (CH3C00Na-3H20) and 14.714 g. of the veronal salt. Five c.c. portions of this solution are treated with 2 c.c. of an 8.5% NaCl solution, with a c.c. 0.1 N HCl, and with (18 — a) c.c. of water. The following table shows how o and pH (hydrogen electrode 25°) are related. 

Fig. 2. Electrophoretic patterns of crystalline papain (153). A, tracings of the electrophoretic patterns of twice recrystallized papain, pH 3.9 protein concentration—1.0% in acetate buffer containing 0.001 M Versene and 0.02 M cysteine. Ascending and descending patterns are shown after 275 minutes of migration. The minor peak evident on the descending pattern represents less than 2% of the total protein and was absent in most of the preparations. B shows the descending pattern at 0.2% protein concentration in Veronal buffer at pH 7.5 after 150 minutes. C is the descending pattern after 150 minutes in acetate buffer at pH 6.0 at a protein concentration of 0.4%. Cysteine and Versene were omitted in B and C. The patterns in B and C are at approximately twice the linear enlargement of those in A.

Fig. 5. Sedimentation constants (iSjo, ) of crystalline papain as a function of protein concentration (153). A shows studies at pH 3.9 to 4.0 in acetate buffer (0.1 to 0.2 ionic strength) at a cysteine concentration of 0.02 M. Results with ( ) added Versene at 0.002 M and without (O) Versene. B shows results in acetate at pH 4.0 (0.1 ionic strength) in the absence of cysteine ( ) and for papain oxidized with iodine (O). C gives measurements with 0.02 M cysteine at 0.1 ionic strength in acetate at pH 5.4 (O) and in Veronal at pH 7.0 ( ). For B and C the straight lines have arbitrarily been drawn to an extrapolated value of 2.42 S for Sia,u at zero concentration this is the average value found at pH 4 in the presence of cysteine.

Figure 7.7 Zeta potentials (calculated from electrophoretic mobility data) relating to particles of different ionogenic character plotted as a function of pH in acetate-veronal buffer at constant ionic strength of 0.05 mol dm 3, (a) Hydrocarbon oil droplets, (b) Sulphonated polystyrene latex particles, (c) Arabic acid (carboxylated polymer) adsorbed on to oil droplets, (d) Serum albumin adsorbed on to oil droplets...

The mobile polar phase was an aqueous buffer (sodium acetate-Veronal buffer 1/7M at pH 7.4) alone or mixed with various quantities of acetone. The compounds were dissolved in water, acetone, or ethanol (1—3 mg/ml) and spotted in 1 jaliter amounts. The spots were detected by an alkaline solution of potassium permanganate. The penicillins could be detected also by iodine azide solution. 

Buffers commonly used in clinical and molecular diagnostic laboratories include acetate, boric acid-borate, carbonate-bicarbonate, citrate diethanolamine, glycine-glycinate, phosphate, phthalate, tris(hydroxymethyl) aminomethane, and veronal-sodium veronal, and the so-caUed Good buffers (Table 1-15). 

Arrow indicates the position of start. The patterns were not corrected for electroendosmosis. Electrophoretic conditions 11 X 2 cm. cellulose acetate film, using veronal buffer at pH 8.6 (I = 0.05) 2-hour run at 0-2°C. and at constant current of 0.6 ma./cm. The cellulose activity was assayed by CMC-saccharification and expressed in terms of the absorbancy at 660 n.m. 

For use, mix 291 ml of solution (a) with 709 ml of solution (6), producing a solution of pH 10. (If necessary, adjust to this value.) Normal saline solution 0.9% aqueous solution of sodium chloride. Buffer solution for electrophoresis dissolve 30.0 g of barbital sodium (= Veronal sodium), 20.0 g of sodium acetate trihydrate, and 0.3 g of anhydrous calcium chloride in water, add 180 ml of 0. IN hydrochloric acid solution and then sufficient water to make 4,000 ml of solution pH = 8.6 r = 0.06. 

Polar mobile phase sodium acetate-Veronal buffer (pH 7.0) containing 2% acetone and satd. with the silicone oil Detection... 

Fig. 3. Electrophoretic mobility as a function of pH for crystalline papain (153). The runs were made at 1.5° in univalent buffers at 0.1 ionic strength. Ac. is acetate, V is Veronal, and G is glycine.

The electrophoretic mobility in acetate buffo , pH 5.0, and in veronal buffer, pH 8.6, was observed to be —15.8, and —i8.6-io cm /sec volt respectively. The molecular weight is small, since the substance dialyzed slowly through Visking membranes values between 1240 and 2075 have been observed for especially low molecular weight, highly polydisperse samples, isolated from patients with Hurler s S3mdrome . ... 

---