Glycine-HCl buffer

OTC, TC, CTC and its isomers in muscle and kidney Extraction with glycine-HCl buffer, cleanup with isolute cyclohexyl cartridge DL = 10 ng/g (muscle) 20 ng/g (kidney) Rec = 56-74% [39]... 

Elute Fc and undigested IgG bound to the immobilized protein A column with 0.1M glycine-HCl buffer, pH 2.8. 

Monoclonal antibodies against STR were used for the preparation of an immunoaffinity chromatography column. Milk samples were defatted by centrifugation and diluted with phosphate-buffered saline. After loading onto the column, this was washed with saline, and STR and DIHS were eluted with the glycine-HCl buffer. The column bounded 80.4% and 88.7% of milk samples containing 100 ppb STR and DIHS, respectively (117). 

Porous Teflon membrane 5. The cell with glycine-HCl buffer (pH 3.1) 6. AgzOi cathode 7. Electrolyte (0.1 M phosphate buffer, pH 7.0) 8. Ammeter 9. Recorder... 

Glycine-HCl buffer O.IM Glycine, pH 2.5. Adjust pH with IMHCl. Sodium acetate buffer 0.06M CHgCOONa, pH 4.6. Adjust pH with acetic acid. 

Blanchflower et al. [43] reported the analysis of TC, oxy-TC, and chlor-TC in pig mnscle and kidney tissne by after extraction in a glycine-HCl buffer and solid-phase extraction (SPE) clean np of the extracts. Gradient LC-APCI-MS was performed with 10-90% acetonitrile in water with 0.04% heptafluorobutyric acid, 10 mmol/1 oxalic acid, and 10 gmol/1 EDTA. Detection limits were 10 gg/kg in muscle and 20 gg/kg in kidney. Ion ratio measurements, e.g., on m/z 410, 426, and 445 for tetracycline, were performed for confirmation. 

In contrast, this method is suitable for APase (Mason and Sammons, 1978). Antibodies to bovine intestinal APase (type VII from Sigma, or type I purified according to Section 10.2.3.3) were produced in the rabbit (Sections 5.3.3.1 and 5.3.4). The enzyme (2.5 mg) was coupled to CNBr-activated Sepharose (0.25 ml Section 7.1.8.2.2) and the specific anti-APase antibodies from 0.5 ml aliquots of rabbit antiserum (or Ig preparation) were retained on such minicolumns which were eluted subsequently with 100 mM glycine-HCl buffer, pH 2.5. The total yield over 7 adsorption/elution cycles is about 1-1.5 mg of antibody per ml of antiserum. In the test, APase and the specific anti-enzyme antibody could then be added simultaneously with the linking antibody (Mason and Sammons, 1978). The specificity of the purified antibody made it also possible to use a crude preparation of the enzyme without any decrease in detectability. Nevertheless, this method is quite involved for routine applications of EIA and is primarily used in EIH, such as in the simultaneous immunoenzymatic detection of two different antigens or epitopes (Section 17.3.2.3). 

Partial denaturation of IgG is carried out with samples containing 10 pg/ml in 50 mM glycine-HCl buffer, pH 2.5, containing 100 mM NaCl, incubated for 10 min at room temperature and neutralized with 500 mM Tris. The sample is then dialyzed against the coating buffer. Alternatively, IgG may be denatured in neutral buffer by the addition of an equal volume of 6 M urea and incubation overnight at room temperature, followed by extensive dialysis. Thermal denaturation is carried out for 10 min at 70°C for sheep antibodies or at 82°C for rabbit antibodies (Conradie et al., 1983). This treatment may be different for IgG preparations from other species. 

The blotted proteins are reacted with their corresponding antibodies (Section 16.3.2). The bands are excised and eluted with 3 ml 200 mM glycine-HCl buffer, pH 2.8, during 2 min at 0 C. The solution is then neutralized with NaOH and concentrated to 0.1-0.2 ml. The monospecific antibodies obtained can then be used for other EIA (blots, titration, EIH). Though the blot can be used several times to extract monospecific antibodies from the serum, the amounts obtained are not excessive. It can be advantageous to link the antigen to the blot by the method given in Table 16.11C. 

Some antigens or epitopes may be hidden, e.g. J-chain in IgA or IgM in ethanol-fixed tissues, which may be exposed after treatment for 1 h at 4°C with 100 mM glycine-HCl buffer, pH 3.2, containing 6 M urea (Brandtzaeg, 1976). 

A similar procedure was used for antidigoxin antibody assay. Digoxin-benzo crown ether conjugates, rather than cortisol were entrapped in the PVC membrane of the K+ sensitive electrode. The detection limit was in the range of a few g/ml. The sensor was regenerated by a brief (<60 s) immersion in glycine-HCl buffer, pH 2.8. The membrane was stable for 1-2 weeks under conditions of routine use. 

Figure 5.15 Equilibrium perturbation. Absorbance changes at 250 nm consequent upon the addition of E. coli purine nucleoside phosphorylase (ISOpg in 5 iL) to a solution (3mL) containing [l - H]inosine (1.99mM), hypo-xanthine (O.lOOmM), ribose-1-phosphate (O.SOmM) and inorganic phosphate (ll.SmM) in 0.2 M glycine-HCl buffer, pH 9.4, in a 1cm pathlength cuvette. ...

Fig. 8. Gel filtration of apoferritin (10 mg) at pH 2.16 in 10 raM glycine/HCl buffer on a column (100 cm x 1 cm) of Sephadex G-100. The peaks, in order of elution, correspond to oligomer, subunit-tetramer, subunit-dimer and subunit (if. if. Crichton, in preparation)...

Sample preparation 10 mL Urine -I- 5 g NaCl + 1 mL water -(- 1 mL concentrated HCl -I- 10 mL Methyl ether, shake for 10 min at 200 oscillations/min, centrifuge at 1540 g for 10 min. Repeat extraction. Combine extracts, add 1 mL 1 M pH 3.5 glycine HCl buffer, shake, centrifuge, iiyect aliquot of lower aqueous phase... 

In another study (114), three different immobilization methods were evaluated for the detection of E. coli using an antibody coated crystal. The best results were obtained with a protein A coated method, and a response for 10 - lO cells 1 ml of E. coli K12 and other antigens of the Enterobacteriaceae family was observed. Repeated usage of the coated crystal by removing the bound antigen with urea or glycine HCl buffer (pH 2.6) was not very successful. 

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