Purification of poly ribosylribotil phosphate capsule from heamophilus influenza type- b strain atf2 cultivated in bioreactor

Mahsa Safa,1,* Farhad esmaily,2 Fatemeh tahoori ,3 Atefeh hadi ,4



Haemophilus influenzae is a small gram negative cocobacilli non motile and can be seen as pleomorphic. h.inf.can cause meny disease like meningitis,pneumonia,epiglottitis,arthritis and cellulitis.it is the main causative agent of meningitis in children under two up to five years. capsule from haemophilus influenzae which is made of poly ribosylribitol phosphate (prp) play a significant role in invasiveness of this organism. haemophilus influenzae type b vaccine (hib) is compulsory and recommended by who as in combination with dtp, hepatitis as dtp,hep,hib.


In this work we extracted capsular polysaccharide (prp) from h.inf. strain atf2 which is isolated from a child with meningitis and cultured in modified gc broth in bioreactor. since polysaccharide antigen are in general small molecules and cannot stimulates the immune t-cells (t-cell independent).the prp has to be conjugated with a carrier protein e.g.; tetanus toxoid (tt) and then the conjugated product (prp-tt) is capable of stimulating the t-cell or it will be t-cell dependent antigen. during the process of conjugation the purified prp has to be activated using cnbr and by the help of adh as spacer it will be attached to tt which is activated by edac . we used sepharose cl-4b gel filtration for separation of conjugated fractions (prp-tt) from free protein after elution of gel. doc was used to measured free prp from prp-tt after use of sepharose cl-4b gel .


the results of above process are very promising. however, more study and investigations are required to achieve optimum conditions.


The aim of study is to achieve the production of hib technology vaccine against meningitis in children.


Haemophilus influenza, conjugation