Simplified purification method for Clostridium difficile toxin A

Figure 1 Purification of Clostridium difficile toxin A by DEAE-Toyopearl 650 mol/L chromatography. Fractions 1-34 were eluted with 50 mmol/L Tris-HCl buffer (pH7.5) containing a linear NaCl gradient (50 mmol/L-250 mmol/L), then fractions 35-50 were eluted with 300 mmol/L NaCl and fractions 51-75 were eluted with linear gradient (300 mmol/L-600 mmol/L NaCl) in the same buffer.

Figure 2 Analysis of toxin A preparations by native-PAGE (4%-30%). Line 1: Purified C. difficile toxin A; Line 2: High molecular mass markers, thyroglobulin (M_r 669000), ferritin (M_r 440000), catalase (M_r 232000), lactate dehydrogenase (M_r 140000), bovine serum albumin (M_r 67000).

Figure 3 Analysis of toxin A by Ouchterlony double immunodiffusion. Well 3 contained 20 μL of antiserum against strain 10463 culture filtrate, well 1 and well 2 contained 20 μL of purified toxin A by acid precipitation and DEAE-Toyopeal 650 mol/L chromatography respectively.