CLIAMS:1. A scalable two-step method for purification of immunogenic diphtheria toxoid, said method comprising the steps of:
a. obtaining an immunogenic diphtheria toxoid containing material prepared from fermentation of Corynebacterium diphtheriae in deferrated papain digest medium;
b. carrying out a first step of separation of diphtheria toxoid using a tangential flow filtration setup having membrane pore size in the range of 100-500kDa to obtain a filtrate;
c. carrying out a second step of separation of filtrate from step b) using a tangential flow filtration setup having membrane pore size in the range of 10-30kDa to obtain a retentate comprising of purified immunogenic diphtheria toxoid,
wherein the purity of diphtheria toxoid is in the range of 1800-2200Lf/mg pN, and wherein the percentage recovery of said purified diphtheria toxoid is in the range of 85-95%.
2. The method as claimed in claim 1, wherein percentage recovery of diphtheria toxoid in said filtrate is in the range of 85-95%.
3. The method as claimed in claim 1, wherein percentage recovery of diphtheria toxoid in said retentate is in the range of 85-95%.
4. A scalable one-step method for purification of immunogenic diphtheria toxoid, said method comprising the steps of:
a. obtaining an immunogenic diphtheria toxoid containing material prepared from semi-synthetic medium;
b. carrying out a separation of diphtheria toxoid using a tangential flow filtration setup having membrane pore size in the range of 10-30kDa to obtain a retentate comprising of purified immunogenic diphtheria toxoid,
wherein the purity of diphtheria toxoid is in the range of 1700-2600Lf/mg pN, and wherein the percentage recovery of said purified diphtheria toxoid is in the range of 85-90%.
5. The method as claimed in any of the claims 1-4, wherein said purified immunogenic diphtheria toxoid is optionally further filter sterilized with a 0.2µm filter.
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