GMP Compliance for Production of CB.Hep-1 Monoclonal Antibody as a Biological Reagent - - BioPharm International

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GMP Compliance for Production of CB.Hep-1 Monoclonal Antibody as a Biological Reagent

Inoculum Preparation

Inoculum preparation includes maintaining the Master and Working Cell Banks (MCB/MWCB) and growing the suspended or stationary methods in spinner flasks. Each production batch is originated from a fresh ampoule of the seed (e.g., working cell bank). The maximum permissible number of serial passages in culture during normal production is defined and restricted. Justification of this limit should include information concerning the yield of monoclonal antibodies and the stability of the hybridoma.

Subsequently, 106 hybridoma cells are inoculated intraperitoneally in each mineral oil primed BALB/c mice.8 Media and buffer preparation is performed manually with powders in mixing tanks.


Figure 2. Inspection Points. Following international regulation we introduced some inspection points and quality specifications for the MAb CB.Hep-1. These tests were established keeping in mind the results and consistency of the process for about six years.
Purification consists of several chromatography gel filtration cycles in Sephadex G-25 coarse (Amershan-Pharmacia, Uppsala, Sweden) Protein an affinity.9 After this chromatography, incubation in 0.1 molar (M) citric acid pH 3.0 for an hour at 4°C was carried out. The ultra filtration and sterile filtration steps complete the basic diagram of the purification process.10 The purified antibody is stored at 4°C for further use as immunoligand to purify the rHBsAg. The final product of the purification process is controlled and released by the Quality Control Laboratory. (See Figure 2.)

Immunoaffinity Matrix


Figure 3. Quality Specification of the Immunoaffinity Matrix
The purified antibodies are bound to the Sepharose CL-4B (Pharmacia) activated with cyanogen bromide, as recommended by the manufacturer (Pharmacia-LKB, 1993). Before release of the columns, they are submitted to a test to verify their functional performance. (See Figure 3.)

Bio-safety of the Process

In order to guarantee the quality and safety of the immunogel CB.Hep-1, the production process is submitted to a strict control of the biological material involved in the process, which is based on the following strategy:

1. Serologic control of the colonies of mice used to produce the ascitic fluid CB.Hep-1
2. Certification of the fetal bovine serum
3. Certification of the master cell bank
4. Virological control of the ascites
5. Viral validation of the purification process

Alemán R and Valdez R, two of our principal investigators in two separate works, have published these strategies.10,11


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