Recombinant human interleukin-2 (rhIL-2) is a recombinant protein product, a non-glycosylated protein with the same biological activity as native IL-2, containing 133 amino acid residues and a relative molecular mass of 15.5 k. Its pharmacological action is to enhance the immune response. Recombinant human IL-2. Approved by the FDA in the United States in 1992 under the trade name Proeukin. Since then, it has been listed in Japan, Denmark and other countries, and the approved clinical application indications are kidney cancer, angiosarcoma and melanoma.

Pharmacopoeia standard

Definition, composition and use

It is made of recombinant human interleukin-2 obtained by fermentation, isolation and high purification of Escherichia coli which expresses human interleukin-2 (human interleukin-2). Suitable stabilizer, no preservatives and antibiotics

Basic requirements

The production, testing and use of facilities, raw materials and accessories, water, utensils, animals, etc. shall comply with the relevant requirements of the "General".

Manufacture

1 engineering strains

1.1 Name and source
The recombinant human interleukin-2 engineering strain is an E. coli strain transformed with a recombinant plasmid carrying the human interleukin-2 gene.

1.2 Establishment of seed lot
It shall comply with the provisions of the “Management Regulations for the Use of Bacterial Species for the Production of Biological Products”.

1.3 strain test
The main seed batch and the working seed batch should be subjected to the following comprehensive tests.
1.3.1 Sorting LB agar plates
It should be in the form of typical E. coli colonies without the growth of other bacteria.
1.3.2 Staining microscopy
It should be a typical Gram-negative bacillus.
1.3.3 Resistance to antibiotics
Should be consistent with the original species.
1.3.4 Electron microscopy (work seed batch can be exempted)
It should be in the form of a typical E. coli, without mycoplasma, virus-like particles and other microbial contamination.
1.3.5 Biochemical reaction
Should meet the biochemical reaction characteristics of E. coli.
1.3.6 Human interleukin-2 expression
Culture in a shaker should not be lower than the expression level of the original strain.
1.3.7 Plasmid check
The restriction map of this plasmid should be identical to the original recombinant plasmid.
1.3.8 Target gene nucleotide sequence check (work seed batch can be exempted)
The nucleotide sequence of the gene of interest should be consistent with the approved sequence.

2 stock solution

2.1 Preparation of seed liquid
The qualified working seed strains are inoculated in a suitable medium (which may contain an appropriate amount of antibiotics).

2.2 Fermentation medium
Use a suitable antibiotic-free medium.

2.3 Seed liquid inoculation and fermentation culture
2.2.3.1 Inoculate an appropriate amount of seed solution in the sterile medium.
2.2.3.2 Fermentation at a suitable temperature shall be carried out according to the approved fermentation process and the corresponding fermentation conditions, such as temperature, pH, dissolved oxygen, feed, fermentation time, etc., shall be determined. The fermentation broth should be checked regularly for plasmid loss rate.

2.4 Fermentation solution treatment
The cells are collected and treated in a suitable manner.

2.5 preliminary purification
Preliminary purification is carried out using an approved purification process to achieve the specified purity.

2.6 Highly purified
After preliminary purification, it is highly purified by an approved purification process to meet the requirements of 3.1, which is a recombinant human leukotriene-2 stock solution. Add a suitable stabilizer, sterilize and filter, store at a suitable temperature, and specify the expiration date.

2.7 stock solution test
According to item 3.1

3 Semi finished products

3.1 Preparation and sterilization
3.1.1 Preparation of diluent
Formulate the dilutions in an approved formulation. It should be used for dilution immediately after preparation.
3.1.2 Dilution and sterilization
The recombinant human interleukin-2 stock solution, which has been tested as qualified plus stabilizer, is diluted to the desired concentration with the dilution of 2.3.1.1. After sterilization and filtration, it is a semi-finished product and stored at 2-8 °C.
3.2 Semi-finished product verification
According to item 3.2.

4 Finished products
4.1 Batch
4.2 Packing
4.3 Specifications
4.4 Packaging

Author's Bio: 

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