Molecular cloning, expression in Escherichia coli, and development of monoclonal antibodies to feline cystatin C.
Cystatin C is a member of the cystatin superfamily of low-molecular-weight proteins that inhibit the activity of cysteine protease. The full-length cDNA that encodes feline cystatin C (FeCysC) has been cloned from feline white blood cells by the oligo-capping method. The cDNA consists of 796 nucleotides and includes an open reading frame encoding a polypeptide of 146 amino acids. Next, we developed several mouse anti-FeCysC monoclonal antibodies (mAbs). The recombinant FeCysC (rFeCysC) was expressed in Escherichia coli (E. coli) BL21 and used as an antigen to immunise mice. The reactivity of the mAbs to native FeCysC was examined by western blot analysis against the urinary protein from cats with chronic kidney disease (CKD). The three anti-rFeCysC mAbs (3-9G, 7-7C, and 9-12F) were able to recognise native FeCysC.
Nakata
Junya
J
Department of Small Animal Internal Medicine, School of Veterinary Medicine and Animal Science, Kitasato University, Higashi-23-35-1, Towada-shi, Aomori 034, Japan.
Nakahari
Aiko
A
Takahashi
Chika
C
Chikazawa
Seishiro
S
Hori
Yasutomo
Y
Kanai
Kazutaka
K
Ito
Naoyuki
N
Higuchi
Seiichi
S
Hoshi
Fumio
F
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D001483:Base Sequence; D002415:Cats / Q000235:genetics* / Q000276:immunology*; D003001:Cloning, Molecular; D055316:Cystatin C / Q000235:genetics* / Q000276:immunology*; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D051379:Mice; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D017386:Sequence Homology, Amino Acid
D000911:Antibodies, Monoclonal; D055316:Cystatin C; D017931:DNA Primers; D018076:DNA, Complementary; D011994:Recombinant Proteins