Production of active anti-CD6 mouse/human chimeric antibodies in the milk of transgenic mice.
The expression of chimeric genes in the mammary gland of transgenic farm animals has become an alternative for the large-scale production of recombinant proteins and for the modification of milk composition. In this paper, we show that a mouse/human chimeric antibody against the human CD6 leukocyte antigen can be assembled and correctly folded by the mammary gland, and secreted to milk, where it maintains its specificity. The base sequences encoding for the heavy and light chain variable regions of the anti-CD6 mouse monoclonal antibody IOR-T1 were cloned by the polymerase chain reaction from hybridoma cDNA, coupled to human heavy and light chain constant region genes, and inserted in a vector containing the 5' regulatory region of the rabbit whey acidic protein gene. Transgenic mice were produced by conventional pronuclei microinjection techniques. Integration and transgene copy number were determined by Southern blot. Assembled human immunoglobulin was detected in milk using a sandwich ELISA. Expression levels of chimeric antibodies in milk were determined to be around 400 micrograms/ml by Western blot, using CHO-derived chimeric IOR-T1 antibodies as reference. The chimeric antibodies produced in milk recognized human peripheral blood T lymphocytes by indirect immunofluorescence, with the classical patch-like pattern of IOR-T1.
Limonta
J
J
Mammalian Cell Genetics Division, Center for Genetic Engineering and Biotechnology, Havana, Cuba.
Pedraza
A
A
Rodríguez
A
A
Freyre
F M
FM
Barral
A M
AM
Castro
F O
FO
Lleonart
R
R
Gracía
C A
CA
Gavilondo
J V
JV
de la Fuente
J
J
D000818:Animals; D000906:Antibodies / Q000235:genetics*; D015703:Antigens, CD / Q000276:immunology*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000276:immunology*; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008321:Mammary Glands, Animal / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D008892:Milk / Q000737:chemistry*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*
D000906:Antibodies; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; C062279:CD6 antigen; D011993:Recombinant Fusion Proteins