CS252599B1 - Cloned human lymphoma cell line CZAS Jurkat K5-2B6 - Google Patents

Abstract

The solution concerns a cloned line of human lymphoma cells IMG CZAS Jurkat K5-2B6, producing interleukin 2. The line can be used industrially as a source of human interleukin 2 for experimental work in immunological differentiation and other model systems, and can also be used for some areas of medical practice.

Description

252599 2

The invention relates to a cloned line of human lymphoma cells Jurkat K5-2B6 producing interleukin-2 (T-cell growth factor).

Human interleukin-2 can be prepared in a conventional manner where the source of interleukin-2 is the supernatant of human lymphocytes activated by lectins. This method has several disadvantages. The supernatant of these cultures contains a heterogeneous mixture of many interleukins, lymphokines and mediators, the spectrum and titer of which are different in each individual supernatant. In addition, the interleukin 2 prepared by this process has a very fluctuating and, on average, low concentration, under optimal conditions of about 1.0 µ / ml medium.

The above-mentioned shortcomings of the above-mentioned and hitherto used method are omitted when the monoclonal humaninterleukin 2 producing human leukemic cell line Jurkat K5-2B6 is disposed of and deposited in the Collection of Cell Lines of IMG CSAV Prague 6, Flemingovo nám. 2 under IMG CZAS Jurkat K5-2B6. Said line was obtained from the starting material of the non-cloned Jurkat line producing less than 0.1 µ / ml (Kaplan et al., Amer. J. Hematol. 1, 219, 1976) in a manner known from the literature (Farrah et al., J. Immunol., 125, 2565, 1980). A greater number of leukemic lines and sublines were tested for interleukin-2 production, namely both constitutive and lectin-induced production, phorbol myristate acetate and combinations thereof. Of all the lines tested, Jurkat-0 was the most preferred subline for further clonal selection. This subline was repeatedly cloned and a clone with the highest interleukin-2 production, designated Jurkat K5-2B6, was selected from approximately 200 clones. The advantage of the cloned line is that after stimulation with phorbol myristate acetate and lectin (phyto-hemagglutinin) it produces a homogeneous cell product - interleukin 2 - in a relatively high concentration. This product has an immunoregulatory effect and can serve to enhance the response in the body. The cloned Jurkat K5-2B6 cell line can be cultured in vitro in suitable animal cells. From canned foods stored in liquid nitrogen, the production of interleukin 2 can be initiated after the cells have been grown to an appropriate level and the phorbol myristate acetate and phyto-hemagglutinin added. Interleukin 2 produced by the cloned high-production line Jurkat K5-2B6 specifically stimulates the synthesis of ONK mammalian (human, mouse, rat) T cells in vitro and mimics the immune response mediated by these lymphocytes. The monoclonal product has a concentration of 10-60 units / ml and it is not necessary to dispose of ballast mediators, lymphokines and interleukins. Example

Jurkat K5-2B6 cells were cultured at 2.10 ® cells / ml for 24 hours in tissue culture containing 50 µg / ml phorbol myristate acetate and 1.5 µg / ml phytohemagglutinin in a total volume of 200 ml medium. After the culture was complete, the culture medium was cleared of cells and filtered. A total of 200 ml of culture medium was obtained which contained 40 units of interleukin 2 per ml, i.e. 8,000 units of interleukin 2 in total. This amount corresponds to conventional Interleukin 2, which can be obtained from 8,000 ml of human peripheral blood in a conventional method of preparation, and only under optimal conditions. Interleukin 2 reacted with the CTLL immune cell line (Gillis and Smith, Nature 288, 154, 1977), which specifically corresponds to DNA synthesis only to interleukin 2. DNA synthesis can be quantitated by the incorporation of HH-TdR; thisincorporation is directly proportional to the concentration of interleukin 2. The CTLL line as well as other line-immune lymphocytes can be infinitely propagated in interleukin 2, while without interleukin 2 such cells die within 24 hours. Tissue-propagated immune lymphocytes can then be used to inject into an organism that lacks such cells, thereby enhancing an immune response against a foreign anti-gene or a tumor of its own, or compensating for immunodeficiency states. Cells of the cloned lineage K5-2B6 grow as suspension or semi-suspension cultures. The basic culture medium is RPMI 1 640 supplemented with non-essential amino acids, L-glutamine, penicillin, streptomycin and 10% inactivated bovine serum (Bioveta, Ivanovice na Hané).

Jurkat K5-2B6 can be industrially used as a source of human interleukin 2 for experimental work in immunological, differentiation, and other modeling systems, and may be useful in some areas of medical practice in the future.

Claims (2)

1 252599 OBJECT OF THE INVENTION The cloned human lymphoma cell line, CZAS Jurkat 2 K5-2B6, producing interleukin.