LIT212C 6/17/98 12:25 PM Page FC1 AG® 1, AG MP-1 and AG 2 Strong Anion Exchange Resin Instruction Manual
LIT212C 6/17/98 12:25 PM Page FC2 Table of Contents Page Section 1 Introduction ............................................ 1 Section 2 Technical Description............................. 1 Section 3 Mechanism .............................................. 5 Section 4 Resin Conversion .................................... 10 Section 5 Instructions for Use ................................ 14 5.1 5.2 Batch Method.................................................... 14 Column Method .................
LIT212C 6/17/98 12:25 PM Page 1 Section 1 Introduction AG 1, AG MP-1 and AG 2 resins are strongly basic anion exchangers. They are capable of exchanging anions of acidic, basic, and neutral salts, and ampholytes on the basic side of their pI. Strong anion exchange resins are used for sample preparation, enzyme assays, metal separations, and peptide, protein, and nucleic acid separations.
LIT212C 6/17/98 12:25 PM Page 2 AG 1 and AG 2 resins are strongly basic anion exchangers with quaternary ammonium functional groups attached to the styrene divinylbenzene copolymer lattice. The amount of resin crosslinkage determines the bead pore size. A resin with a lower percentage of crosslinkage has a more open structure permeable to higher molecular weight substances than a highly crosslinked resin.
LIT212C 6/17/98 12:25 PM Page 4 Each AG 1 resin is supplied in the chloride form. Selected resins are available in the acetate, formate, and hydroxide form. These ionic forms may be considered more activated forms than the chloride form, as may be deduced from the order of selectivity information given in Tables 2 and 3. AG 1 resins purchased in the more active forms may be converted to any other form.
LIT212C 6/17/98 12:25 PM Page 6 AG MP-1, neutral species and cations do not interact with the resin. In the chloride form of AG 1, AG MP-1, and AG 2 resin, the counterion on the resin is Cl -. A resin can be converted from one ionic form to another. Usually the resin is used in an ionic form with a lower selectivity for the functional group than the sample ions to be exchanged.
LIT212C 6/17/98 12:25 PM Page 8 The AG 1 and AG MP-1 resins are available in several particle size ranges. The flow rate in a chromatographic column increases with increasing particle size. However, the attainable resolution increases with decreasing particle size and narrower size distribution ranges. Particle size is given either in mesh size or micron size. The larger the mesh size number, the smaller the particle size. Table 4 shows wet mesh and equivalent micron diameters.
LIT212C 6/17/98 12:25 PM Page 10 Section 4 Resin Conversion Table 5 outlines common techniques for converting ion exchange resins from one ionic form to another. Resin conversion is most efficiently carried out in the column mode. However, when choosing a column, remember that the resin may shrink, or it may swell as much as 100%, depending on the conversion. Table 5.
LIT212C 6/17/98 12:25 PM Page 12 Conversions to ionic forms not listed in Table 5 can be achieved using the information supplied in Table 3, which lists relative selectivities of various counterions for AG 1, AG MP-1, and AG 2 resin. To convert a resin to an ionic form with a higher selectivity, wash the resin with 2-5 bed volumes of a 1 M solution of the desired counterion.
LIT212C 6/17/98 12:25 PM Page 14 Section 5 Instructions for Use AG 1 and AG MP-1 resins may be used in a batch method or a column method. The batch method consists of adding the resin directly to the sample and stirring. The column method requires packing a column with resin, and passing the sample through. 5.1 Batch Method The batch method is performed by adding the resin directly into the sample and stirring. The resin should be in the correct ionic form prior to beginning. 1.
LIT212C 6/17/98 12:25 PM Page 16 exchanged onto the column. For unknown solutions, use deionized water. 4. Slurry and pour the resin into the column. Equilibrate the resin in the initial buffer using 3 bed volumes of buffer. Poorly equilibrated resin will not give reproducible results. Alternatively, equilibration can be done by the batch technique, prior to pouring the column. First, convert the resin to the appropriate form, then suspend it in the starting buffer.
LIT212C 6/17/98 12:25 PM Page 18 Table 6. Suggested Flow Rates for Ion Exchange Resin Columns Application Removing trace ions Separations with very few components Separations of multi-component samples Using high resolution resins with small particle size Linear Flow Rate (cm/min) Materials AG 1-X8 resin, 50-100 mesh, 10 grams 5-10 1-3 0.3-1.0 Glass column approximately 12 mm ID, 30-40 cm long, and resistant to 9 N HCl 1-2 Sample solution - 0.
LIT212C 6/17/98 12:25 PM Page 20 3. Equilibrate the resin by passing approximately 15 ml of 9 N HCl over the resin bed. 4. Adjust the flow rate to no faster than 1 drop/second or 4 ml/min. 5. When the level of 9 N HCl has reached the top of the resin bed, either shut the screw clamp, or add 2 ml of sample (0.2 meq of each ion). 6. Do not allow air into the resin bed because air may cause channeling with uneven flow of subsequent solutions. 7.
LIT212C 6/17/98 12:25 PM Page 22 Ames Keta-Diastix test for glucose 1 M NaCl Procedure 1. Pack the Poly-Prep chromatography column with 2 ml AG 1-X8 resin, 200-400 mesh, formate form. 2. Convert the resin to the OH form by washing it with 10 bed volumes of 1 N NaOh, then with 5 bed volumes of distilled water. 3. Add 1 ml of the glucose sample. 4. Wash with 3 bed volumes of water. 5. Check for the presence of glucose. 6. Elute the glucose with 1 M NaCl. 7. Check for glucose.
LIT212C 6/17/98 12:25 PM Page 24 Application Resin Reference Application Resin Reference Purification of carboxylated pepsinogen AG 1-X8 resin Rajagopalan, T. G., Moore, S. and Stein, W. J., J. Biol. Chem., 241, 4940, (1966). Removal of thyroid hormone from serum AG 1-X8 resin Stanley, F., Tsai, J. R. and Samuels, H. H., J. Biol. Chem., 261, 9400 (1966). Separation of cAMP from cGMP AG 1-X8 resin Kuehl, F. A., Jr., Ham, E. A. and Zanetti, M. E, et al., Proc. Nat. Acad. Sci.
LIT212C 6/17/98 12:25 PM Page 26 Metals Recommended Eluant and Resin Eluted Ions Ni, Co, Cu, AG 1-X8 Zn resin Reference Ni - 96% MeOH, Fritz, J. S. Pietrzyk, 0.2 M HCl; Co - D. J., Talanta, 8, 143 55% IPA, 1.3 M (1961). HCl; Cu - 55% IPA, 0.1 M HCl; Zn - 0.005 M HCl Mn, Co, Ni AG 1-X8 Fe, Mo, resin (also Cr, Zn, Cd, Hg) Mn, Co, Ni - 8.5 x 10-2 M tartrate; Fe - tartaric acid in 0.1 M HCl; Mo - 3 M NaOH Morie, G. P., and Sweet, T. R., J. Chromatog., 16, 201 (1964).
LIT212C 6/17/98 12:25 PM Page 28 Table 10. Anion Exchange Resins in Enzymatic Assays Enzyme Substrate Product Resin Reference NADase NAD Nicotinamide AG 1-X2 resin Moss, J., Manganiello, V. C. and Vaughn, M., Proc. Nat. Acad. Sci. USA, 73, 4424 (1976). Cyclic 3',5' nucleotide phosphodiesterase cAMP Adenosine AG 1-X2 resin Brooker, G., Thomas, L. J., Jr. and Appelman, M. M., Biochem., 12, 4177 (1968); Ong, K. K. and Rennie, P. I. C., Anal. Biochem., 76, 53 (1976); Thompson, W. J., Teraski, W.
LIT212C 6/17/98 12:25 PM Page 30 Table 10. Anion Exchange Resins in Enzymatic Assays (continued) Enzyme Substrate Hexokinase Mannose Choline kinase ACh + ATP HMG-CoA reductase Glutamine synthetase Resin Reference AG 1-X8 resin Li, E., Jabas, I. and Kornfeld, S., J. Biol. Chem., 253, 7762 (1978). Phosphorylcholine AG 1-X8 resin Kato, A. C., Collier, B. Ilson, D. and Wright, J. M., Can. J. Physiol. Pharmacol., 53, 1050 (1975). HMG-CoA Mevalonolactone AG 1-X8 resin Edwards, P. A.
LIT212C 6/17/98 12:25 PM Page 32 Section 8 Product Information Catalog Number Ionic Form Dry Mesh Size Wet bead Diameter (µm) Capacity (meq/ml) Pkg. Size Nominal Density (gm/ml) AG 1-X2 Resin, Analytical Grade 140-1231 Chloride 50-100 180-500 0.6 500 g 0.65 140-1241 Chloride 100-200 106-250 0.6 500 g 0.65 140-1251 Chloride 200-400 75-180 0.6 500 g 0.65 140-1253 Acetate 200-400 75-180 0.6 500 g 0.65 AG 1-X4 Resin, Analytical Grade 140-1331 Chloride 50-100 180-425 1.
LIT212C 6/17/98 12:25 PM Page 34 Catalog Number Ionic Form Dry Mesh Size Wet bead Diameter (µm) Capacity (meq/ml) Pkg. Size Nominal Density (gm/ml) AG 1-X8 Resin, Analytical Grade (cont.) 140-1431 Chloride 50-100 180-425 1.2 500 g 0.75 140-1441 Chloride 100-200 106-180 1.2 500 g 0.75 140-1443 Acetate 100-200 106-180 1.2 500 g 0.75 140-1444 Formate 100-200 105-180 1.2 500 g 0.75 140-1451 Chloride 200-400 45-106 1.2 500 g 0.75 140-1453 Acetate 200-400 45-106 1.
LIT212C 6/17/98 12:25 PM Page 36 Catalog Number Ionic Form Dry Mesh Size Diameter (micron) Capacity (meq/ml) Pkg. Size Density (gm/ml) AG 2-X8 Resin, Analytical Grade 140-2421 Chloride 20-50 300-1,180 1.2 500 g 0.75 140-2441 Chloride 100-200 90-250 1.2 500 g 0.75 140-2451 Chloride 200-400 45-106 1.2 500 g 0.75 75-180 0.6 100 g 0.65 106-250 1.0 100 g 0.70 106-180 1.2 100 g 0.75 45-106 1.2 100 g 0.
LIT212C 6/17/98 12:25 PM Page 38 Section 9 Technical Information If you need additional technical assistance in using ion exchange resins, contact your local Bio-Rad representative.
LIT212C 6/17/98 12:25 PM Page 39 Bio-Rad Laboratories, 2000 Alfred Nobel Drive, Hercules, CA 94547 LIT212 Rev C
