Characterization of monoclonal antibodies specific to the transcription factor ETS-2 protein

We present the generation of a panel of monoclonal antibodies (F55A10, F55A12, F64A6B4, and F65A2) against the homeodomain transcription factor Nkx6.1, one of the essential transcription factors that regulates the multistep differentiation process of precursor cells into endocrine β-cells in the pancreas. Expression of Nkx6.1 can be detected in developing pancreatic epithelium and in adult insulin-producing β-cells, making this transcription factor a unique β-cell marker. For production of monoclonal antibodies, RBF mice were immunized with a GST-Nkx6.1 fusion protein containing a 66-amino acid C-terminal fragment of rat Nkx6.1. Four clones were established as stable hybridoma cell lines and the produced antibodies were of the mouse IgG1/κ subtype. When applied for immunohistochemistry on frozen sections of adult mouse pancreas, monoclonal antibodies stain specifically the β-cells in the endocrine islets of Langerhans with patterns comparable to that of a previously produced polyclonal rabbit serum. Monoclonal antibodies can be divided into two groups that appear to recognize different epitopes, as determined by competition ELISA. The presented antibodies are useful tools for the further characterization of the role and function of Nkx6.1 in pancreatic development, especially for use in double-labeling experiments with existing polyclonal rabbit antibodies. (J Histochem Cytochem 54:567-574, 2006)

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Generation and Characterization of Monoclonal Antibodies against the ERGB/FLI-1 Transcription Factor

Hybridoma ◽

10.1089/hyb.1995.14.563 ◽

1995 ◽

Vol 14 (6) ◽

pp. 563-569 ◽

Cited By ~ 13

Author(s):

XIAN-KUI ZHANG ◽

TAKIS S. PAPAS ◽

NARAYAN K. BHAT ◽

DENNIS K. WATSON

Keyword(s):

Transcription Factor ◽

Monoclonal Antibodies

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Characterization of Monoclonal Anti-human Factor VII Antibody (B101/B1) that Recognized Three-dimensional Structures near Arg at Position 79 in the First EGF-like Domain

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614229 ◽

1998 ◽

Vol 79 (01) ◽

pp. 104-109 ◽

Cited By ~ 6

Author(s):

Osamu Takamiya

Keyword(s):

Monoclonal Antibodies ◽

Tissue Factor ◽

Human Factor ◽

Solid Phase ◽

Three Dimensional ◽

Coagulation Factors ◽

Factor Vii ◽

Dimensional Structure ◽

Epidermal Growth

SummaryMurine monoclonal antibodies (designated hVII-B101/B1, hVIIDC2/D4 and hVII-DC6/3D8) directed against human factor VII (FVII) were prepared and characterized, with more extensive characterization of hVII-B101/B1 that did not bind reduced FVIIa. The immunoglobulin of the three monoclonal antibodies consisted of IgG1. These antibodies did not inhibit procoagulant activities of other vitamin K-dependent coagulation factors except FVII and did not cross-react with proteins in the immunoblotting test. hVII-DC2/D4 recognized the light chain after reduction of FVIIa with 2-mercaptoethanol, and hVIIDC6/3D8 the heavy chain. hVII-B101/B1 bound FVII without Ca2+, and possessed stronger affinity for FVII in the presence of Ca2+. The Kd for hVII-B101/B1 to FVII was 1.75 x 10–10 M in the presence of 5 mM CaCl2. The antibody inhibited the binding of FVII to tissue factor in the presence of Ca2+. hVII-B101/B1 also inhibited the activation of FX by the complex of FVIIa and tissue factor in the presence of Ca2+. Furthermore, immunoblotting revealed that hVII-B101/B1 reacted with non-reduced γ-carboxyglutaminic acid (Gla)-domainless-FVII and/or FVIIa. hVII-B101/B1 showed a similar pattern to that of non-reduced proteolytic fragments of FVII by trypsin with hVII-DC2/D4 on immunoblotting test. hVII-B101/B1 reacted differently with the FVII from the dysfunctional FVII variant, FVII Shinjo, which has a substitution of Gln for Arg at residue 79 in the first epidermal growth factor (1st EGF)-like domain (Takamiya O, et al. Haemosta 25, 89-97,1995) compared with normal FVII, when used as a solid phase-antibody for ELISA by the sandwich method. hVII-B101/B1 did not react with a series of short peptide sequences near position 79 in the first EGF-like domain on the solid-phase support for epitope scanning. These results suggested that the specific epitope of the antibody, hVII-B101/B1, was located in the three-dimensional structure near position 79 in the first EGF-like domain of human FVII.

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Antigenic characterization of influenza A virus matrix protein with monoclonal antibodies.

Journal of Virology ◽

10.1128/jvi.49.1.248-252.1984 ◽

1984 ◽

Vol 49 (1) ◽

pp. 248-252 ◽

Cited By ~ 26

Author(s):

K L van Wyke ◽

J W Yewdell ◽

L J Reck ◽

B R Murphy

Keyword(s):

Monoclonal Antibodies ◽

Influenza A Virus ◽

Influenza A ◽

Matrix Protein ◽

Antigenic Characterization

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Mating Type in Chlamydomonas Is Specified by mid, the Minus-Dominance Gene

Genetics ◽

10.1093/genetics/146.3.859 ◽

1997 ◽

Vol 146 (3) ◽

pp. 859-869 ◽

Cited By ~ 2

Author(s):

Patrick J Ferris ◽

Ursula W Goodenough

Keyword(s):

Transcription Factor ◽

Mating Type ◽

Codon Bias ◽

Leucine Zipper ◽

Laboratory Strain ◽

Leucine Zipper Motif ◽

Type Locus ◽

Diploid Cells ◽

Necessary And Sufficient

Diploid cells of Chlamydomonas reinhardtii that are heterozygous at the mating-type locus (mt +/mt –) differentiate as minus gametes, a phenomenon known as minus dominance. We report the cloning and characterization of a gene that is necessary and sufficient to exert this minus dominance over the plus differentiation program. The gene, called mid, is located in the rearranged (R) domain of the mt – locus, and has duplicated and transposed to an autosome in a laboratory strain. The imp11 mt – mutant, which differentiates as a fusion-incompetent plus gamete, carries a point mutation in mid. Like the fus1 gene in the mt + locus, mid displays low codon bias compared with other nuclear genes. The mid sequence carries a putative leucine zipper motif, suggesting that it functions as a transcription factor to switch on the minus program and switch off the plus program of gametic differentiation. This is the first sex-determination gene to be characterized in a green organism.

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