scholarly journals Immunolocalization of Aldolase A Subunit Using Monoclonal Antibody in Rabbit Tissues.

1997 ◽  
Vol 30 (5/6) ◽  
pp. 601-608 ◽  
Author(s):  
Shuji Yamashita ◽  
Takashi Sogo ◽  
Kenjiro Yasuda
Keyword(s):  
Monoclonal Antibody ◽  
A Subunit ◽  
Aldolase A ◽  
Rabbit Tissues

Single Step Purification of Platelet Factor XIII Using an Immobilized Factor XIII A-Subunit Monoclonal Antibody

1993 ◽  
Vol 69 (04) ◽  
pp. 397-400 ◽  
Author(s):  
Daniela Lukacova ◽  
Guy L Reed
Keyword(s):  
Monoclonal Antibody ◽  
Factor Xiii ◽  
Enzyme Assay ◽  
Single Step ◽  
Platelet Lysate ◽  
Catalytic Subunit ◽  
Platelet Factor ◽  
Polyacrylamide Electrophoresis ◽  
Immunoaffinity Purification ◽  
A Subunit

SummaryTo facilitate the analysis of the catalytic subunit of factor XIII we have developed a method for the immunoaffinity purification of this protein from platelets. This method employs a monoclonal antibody that binds to the a-subunit of factor XIII. The anti-factor XIII antibody was immobilized on agarose and then incubated with platelet lysate. Subsequently factor XIII was isolated from the platelet lysate in a single step with a 41% yield as measured by enzyme assay. The purified platelet factor XIII appeared nearly homogeneous when analyzed by polyacrylamide electrophoresis and by immunoblotting with another factor XIII monoclonal antibody.

scholarly journals Monoclonal Antibody against the Plasmodium falciparum Chitinase, PfCHT1, Recognizes a Malaria Transmission-Blocking Epitope in Plasmodium gallinaceum Ookinetes Unrelated to the Chitinase PgCHT1

2002 ◽  
Vol 70 (3) ◽  
pp. 1581-1590 ◽  
Author(s):  
Rebecca C. Langer ◽  
Fengwu Li ◽  
Vsevolod Popov ◽  
Alexander Kurosky ◽  
Joseph M. Vinetz
Keyword(s):  
Monoclonal Antibody ◽  
Plasmodium Falciparum ◽  
Synthetic Peptide ◽  
Transmission Blocking ◽  
Reducing Conditions ◽  
Membrane Feeding ◽  
Epithelial Cell Surface ◽  
A Subunit ◽  
35 Kda Protein ◽  
Homologous Amino Acid

ABSTRACT To initiate invasion of the mosquito midgut, Plasmodium ookinetes secrete chitinases that are necessary to cross the chitin-containing peritrophic matrix en route to invading the epithelial cell surface. To investigate chitinases as potential immunological targets of blocking malaria parasite transmission to mosquitoes, a monoclonal antibody (MAb) was identified that neutralized the enzymatic activity of the sole chitinase of Plasmodium falciparum, PfCHT1, identified to date. This MAb, designated 1C3, previously shown to react with an apical structure of P. falciparum ookinetes, also reacts with a discrete apical structure of P. gallinaceum ookinetes. In membrane feeding assays, MAb 1C3 markedly inhibited P. gallinaceum oocyst development in mosquito midguts. MAb 1C3 affinity isolated an ∼210-kDa antigen which, under reducing conditions, became a 35-kDa antigen. This isolated 35-kDa protein cross-reacted with an antiserum raised against a synthetic peptide derived from the P. gallinaceum chitinase active site, PgCHT1, even though MAb 1C3 did not recognize native or recombinant PgCHT1 on Western blot. Therefore, this affinity-purified 35-kDa antigen appears similar to a previously identified protein, PgCHT2, a putative second chitinase of P. gallinaceum. Epitope mapping indicated MAb 1C3 recognized a region of PfCHT1 that diverges from a homologous amino acid sequence conserved within sequenced chitinases of P. berghei, P. yoelii, and P. gallinaceum (PgCHT1). A synthetic peptide derived from the mapped 1C3 epitope may be useful as a component of a subunit transmission-blocking vaccine.

Role of complex asparagine-linked oligosaccharides in the expression of a functional thyrotropin receptor

2001 ◽  
Vol 354 (2) ◽  
pp. 331-336
Author(s):  
Sandrine SIFFROI-FERNANDEZ ◽  
Sabine COSTAGLIOLA ◽  
Sophie PAUMEL ◽  
Annie GIRAUD ◽  
J. Paul BANGA ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Cell Surface ◽  
Molecular Mass ◽  
Cell Line ◽  
Ovary Cell ◽  
Thyrotropin Receptor ◽  
Complex Type ◽  
C Peptide ◽  
A Subunit

To evaluate the functional role of complex asparagine-linked oligosaccharides of the human thyrotropin receptor (TSHR), a Chinese hamster ovary cell line (JP09) and a K562 cell line (K562-TSHR) expressing this receptor were treated with deoxymannojirimycin (dMM), a mannosidase I inhibitor. dMM blocks the formation of complex-type structures and leads to the formation of high-mannose-type structures. Treatment of cells with dMM led to a decrease in the number of thyrotropin (TSH)-binding sites at the cell surface. Detection of the TSHR at the cell surface using a monoclonal antibody directed against the A subunit showed that this decrease was not due to a decrease in the number of TSHRs expressed at the cell surface. However the recognition of TSHR by a monoclonal antibody directed against the C peptide was greatly decreased. On immunoblotting, after deglycosylation using peptide N-glycanase F, the A subunit was visualized as a doublet (36 and 41kDa). In control cells the species of higher molecular mass was more abundant whereas after dMM treatment the species of lower molecular mass became more abundant. This difference in molecular mass between the two peptides is compatible with the removal of the C peptide. In conclusion, the results show that inhibition of complex-type structure formation leads to (i) an incapacity for TSHR to bind TSH, without affecting its intracellular transport and (ii) an increase of TSHR susceptibility to proteases that remove the C peptide. We then hypothesized that removal of the C peptide could contribute to the formation of a non-functional TSHR.

scholarly journals Identification, by a monoclonal antibody, of a 53-kD protein associated with a tubulo-vesicular compartment at the cis-side of the Golgi apparatus.

1988 ◽  
Vol 107 (5) ◽  
pp. 1643-1653 ◽  
Author(s):  
A Schweizer ◽  
J A Fransen ◽  
T Bächi ◽  
L Ginsel ◽  
H P Hauri
Keyword(s):  
Monoclonal Antibody ◽  
Golgi Apparatus ◽  
Adenocarcinoma Cell Line ◽  
Transport Pathway ◽  
Adenocarcinoma Cell ◽  
Transmembrane Topology ◽  
Golgi Membranes ◽  
A Subunit ◽  
Vesicular Compartment ◽  
Subunit Size

Purified Golgi membranes of the human intestinal adenocarcinoma cell line Caco-2 were used as an antigen to produce a monoclonal antibody, G1/93, which specifically labels a tubulovesicular compartment near the cis side of the Golgi apparatus, including the first cis-cisterna itself, as visualized by single and double immunoelectron microscopy with antibodies against galactosyltransferase. The antigen recognized by G1/93 was identified as a protein with a subunit size of 53 kD. Pulse-chase experiments revealed that the 53-kD protein dimerizes immediately after synthesis followed by formation of oligomers of approximately 310 kD, probably homohexamers. The protein has a transmembrane topology with only a short cytoplasmic segment as assessed by protease protection experiments. Glycosidase digestion studies indicated that the protein is probably not glycosylated. The unique subcellular distribution of the G1/93 antigen in close vicinity to the cis-Golgi is in line with the notion that this protein may delineate the biosynthetic transport pathway from the endoplasmic reticulum to the Golgi apparatus. Moreover, G1/93 is a useful marker to identify the cis side of the Golgi apparatus in a variety of human cells.

scholarly journals Oral Intoxication of Mice with Shiga Toxin Type 2a (Stx2a) and Protection by Anti-Stx2a Monoclonal Antibody 11E10

2013 ◽  
Vol 82 (3) ◽  
pp. 1213-1221 ◽  
Author(s):  
L. M. Russo ◽  
A. R. Melton-Celsa ◽  
M. A. Smith ◽  
M. J. Smith ◽  
A. D. O'Brien
Keyword(s):  
Monoclonal Antibody ◽  
Shiga Toxin ◽  
Biological Activities ◽  
Lethal Dose ◽  
Content Type ◽  
Tubular Necrosis ◽  
Food Borne ◽  
Uremic Syndrome ◽  
A Subunit ◽  
Full Protection

ABSTRACTShiga toxin (Stx)-producingEscherichia coli(STEC) strains cause food-borne outbreaks of hemorrhagic colitis and, less commonly, a serious kidney-damaging sequela called the hemolytic uremic syndrome (HUS). Stx, the primary virulence factor expressed by STEC, is an AB5toxin with two antigenically distinct forms, Stx1a and Stx2a. Although both toxins have similar biological activities, Stx2a is more frequently produced by STEC strains that cause HUS than is Stx1a. Here we asked whether Stx1a and Stx2a act differently when delivered orally by gavage. We found that Stx2a had a 50% lethal dose (LD50) of 2.9 μg, but no morbidity occurred after oral intoxication with up to 157 μg of Stx1a. We also compared several biochemical and histological parameters in mice intoxicated orally versus intraperitoneally with Stx2a. We discovered that both intoxication routes caused similar increases in serum creatinine and blood urea nitrogen, indicative of kidney damage, as well as electrolyte imbalances and weight loss in the animals. Furthermore, kidney sections from Stx2a-intoxicated mice revealed multifocal, acute tubular necrosis (ATN). Of particular note, we detected Stx2a in kidney sections from orally intoxicated mice in the same region as the epithelial cell type in which ATN was detected. Lastly, we showed reduced renal damage, as determined by renal biomarkers and histopathology, and full protection of orally intoxicated mice with monoclonal antibody (MAb) 11E10 directed against the toxin A subunit; conversely, an irrelevant MAb had no therapeutic effect. Orally intoxicated mice could be rescued by MAb 11E10 6 h but not 24 h after Stx2a delivery.

A monoclonal antibody recognizes a human nuclear protein resembling Xenopus oocyte nucleoplasmin

1987 ◽  
Vol 87 (5) ◽  
pp. 713-722
Author(s):  
J. Lord ◽  
G. Brown ◽  
C. Bunce ◽  
P. Nathan
Keyword(s):  
Monoclonal Antibody ◽  
Xenopus Oocytes ◽  
Nuclear Protein ◽  
Chinese Hamster ◽  
Nucleosome Assembly ◽  
3T3 Cells ◽  
Nucleosome Assembly Protein ◽  
Subunit Molecular Weight ◽  
Interphase Cells ◽  
A Subunit

The monoclonal antibody 10BG2 (IgG3) was derived from a mouse immunized with human pre B cells. In immunofluorescence studies the antibody revealed a human nuclear-associated determinant, which in interphase cells was entirely restricted to the nucleus. In metaphase cells 10BG2 antigen was detected throughout the cytoplasm with intensified staining at the periphery of chromosomes. 10BG2 antibody stained all human normal and transformed cells tested. In contrast, the antibody did not stain mouse 3T3 cells or Chinese hamster ovary (CHO) cells. Electrophoresis under denaturing and non-denaturing conditions revealed the 10BG2 antigen to be a 130 X 10(3) to 140 X 10(3) Mr protein with a subunit molecular weight of 29.5 X 10(3). This suggests the protein is at least a tetramer. On two-dimensional gels the 10BG2 antigen had a streaked appearance and separated into two isoelectric variants (pI5.2, 5.4). The protein was also shown to be phosphorylated and thermostable, and remained in solution at pH 3.6. 10BG2 antigen was highly soluble in aqueous buffers and co-migrated on non-denaturing gels with the nucleosome-assembly protein, nucleoplasmin, purified from Xenopus oocytes. The similarity of 10BG2 antigen to nucleoplasmin is discussed.

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