rev: January 26, 2005
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ANTIBODIES
(anti-Human and others as indicated)
RDI Divison of researchd Industries Intl offers a wide line of antibodies. Since no one antibody works best for all applications (neutralization, blotting, ELISA, etc), we offer many different types of antibodies to help solve this problem. Please inquire for other applications or types of antibodies not listed below.
ANTI-Endothelin Receptor A & B (see also sample IP and histo protocol below)
-see also new rabbit polyclonals below
cat# RDI-ET1RECPabm $438.00/50ug
RDI-ET1RECPabm-1 $750.00/150ug
Background: Endothelin-1 (ET-1) is a potent vasoconstrictor that is secreted
by vascular endothelial cells. ET-1, ET-2, ET-3, and endothelin b are members
of a family of related peptides. All are 21 amino acid peptides with two
disulfide bridges. ET-1 and ET-2 differ by only two residues, while ET-3
differs from both by six residues. ET-1 and ET-2 are highly effective on
vascular smooth muscle, while the effects of ET-3 are minimal. ET-1 and its
precursor, big ET-1, are the primary forms present in the circulation.
Circulating ET-1 plays a pathologic role in the etiology of hypertension
and antagonists are utilized in the treatment of this disease. The ET-1 receptor
is a seven-transmembrane domain receptor of the G-protein-coupled family
that signals via transient increases in intracellular Ca 2+. It is primarily
expressed in lung and heart and is detected at lower levels in liver, brain,
muscle, and kidney. ET-1 receptor interacts with ET-1, ET-2, and ET-3. However,
it has a 2-3 fold higher affinity for ET-1 than ET-2 and the affinity for
ET-3 is more than 100 fold lower. Thus, ligation of the ET-1 receptor by
ET-1 induces increased intracellular Ca 2+ that signals to promote
vasoconstriction.
Isotype: mIgG1
package: 50ug (200ul) or 150ug (600ul) (at 0.25mg/ml in 50% glycerol, PBS
+ 1.5mM NaN3 & 1mg/ml BSA)
antigen: a peptide protein fragment corresponding to amino acids 230-352
from rat endothelin 1 receptor
USe: Suitable for western blot on human, rat and mouse under using approx
0.5ug/ml ab on total cell of rat testis lysate.(2-3 Hour RT or over night
at 4 DEg C with chemiluminescence detection). Major band =MW 50kDa.
Immunoreactivity also shown with Immunofluorescence
Storage: Store at -20 DEG C upon receipt. . Recommend aliquoting. Avoid frequent
warming cycles.
ref: -Lin, H.Y., et al. 1991 Proc. Natl. Acad. Sci. USA 88:3185- 3189
-Masuda, Y., et al. 1989 FEBS Letters 257:208-210
Precautions: For In vitro research Use Only. Not for use in or on humans
or animals or for diagnostics. It is the responsibility of the user to comply
with all local/state and Federal rules in the use of this product. We are
not responsible for any patent infringements that might result with the use
of or derivation of this product.
DATA SHEET: Sheep anti-Endothelin B Receptor
Catalog#: RDI-ENDTHLBRabS Price: $440.00/250ug vial
Batch: see sheet with shipment
Package Size: packaged at 1mg/ml concnetration in 0.15M PBS, pH 7.5 (no
preservatives)
Antigen: C-terminal peptide of rat endothelin B receptor conjugated to KLH.
sequence: Q T F E E K Q S L E E K Q
Species: Sheep IgG purified by Protein G chromatography
USE: -Western Blot: aprox 20ug/ml (identifies 39Kd band)
-Immunohistochemistry 20ug/ml (parrafin sections).
-immunoprecipitation
Note: These antibodies bind epitopes on the cytoplasmic side of the receptors,
thus the cell membrane must be permeable to antibodies. They will not block binding
of endothelin to the receptors.
Specificity: Tested with endothelin B receptor transfected COS cells. reacts
with human, mouse and sheep.
Storage: Store at 4 DEG C (short term,) -20 DEG C long term. Avoid frequent
freeze thaw cycles.
Precautions: For In vitro research Use Only. Not for use in or on humans
or animals or for diagnostics
Also available: Anti-Endothelin A Receptor cat#RDI-ENDTHLARabS
(same price)
anti-endothelin antibodies (see alphabetical link)
For In Vitro Research Use Only-Not For Use In Or On Humans or Animals
Catalog#: RDI-ENDTHLARabS Price: $440.00/250ug vial
Batch: see sheet with shipment
Package Size: packaged at mg/ml concentration in 0.15M PBS, pH 7.5
(no preservatives)
(VIAL CONTAINS 66UL/250UG).
Antigen: C-terminal peptide of rat endothelin A receptor conjugated to KLH.
Sequence: 410-422 Q E Q N H N T E R S S H K
Species: Sheep IgG purified by Protein G chromatography
USE: -Western Blot: aprox 20ug/ml (identifies 39Kd band)
-Immunohistochemistry 20ug/ml (parrafin sections).
-immunoprecipitation
-Elisa
Note: These antibodies bind epitopes on the cytoplasmic side of the receptors, thus the cell membrane must be permeable to antibodies. They will not block binding of endothelin to the receptors.
Specificity: Tested with endothelin A receptor transfected COS cells. Also
reacts with human, mouse, rat and sheep.
Storage: Store at 4 DEG C (short term,) -20 DEG C long term. Avoid frequent freeze thaw cycles.
Precautions: For In vitro research Use Only. Not for use in or on humans
or animals or for diagnostics
TECHNICAL BULLETIN
For Use With Anti-Endothelin A & Anti-Endothelin B Polyclonal Antibodies.
Techniques For Immunocytochemical Staining & Immunoprecipitation
N.B.: These PAbs recognize epitopes on the cytoplasmic side of the receptors. Thus, to efective use these antibodies, the cell membrane must be permeable to antibodies. These antibodies, therefore, are of significant benefit since they will not block the binding of endothelins to the receptors. (try with and without 0.1% saponin for various tissues)
IMMUNOCYTOCHEMICAL STAINING for ET-A and ET-B receptors
Fixed paraffin-embedded sections (4-6 um) are deparaffinized, stained,
counterstained, dehydrated, cleared and mounted. Block the sections with
10% normal rabbit serum to inhibit non-specific binding. Dilute the Protein
G purified sheep anti-ET-Br or anti-ET-Ar to 10 ug IgG/ml in PBS containing
1.0% normal rabbit serum.Perform the immunoreactivity by using a
peroxidase-coupled rabbit anti-sheep IgG.
IMMUNOPRECIPITATION OF ET-A and ET-B receptors
Prepare cell lysates (-5mg) by homogenizing tissues in 2mL RIPA (50mM Tris, pH = 7.5, 150mMNAC1, 1% NP-40, 0.5% deoxycholic acid,0.1% SDS, 2 mM EDTA, 10 mM NaF, 10 ug/ml leupeptin, 10 ug/ml pepstatin, 20 ug/ml aprotinin). Centrifuge the homogenate for 10 minutes at 1000xg. Dilute 500ul of the cell lysate 1:1 in RIPA buffer. Immunoprecipitate the receptors with 4ug of appropriate antibody for 1 hour at room temperature. Collect the precipitin with Protein G Plus Agarose and wash 3 times with 1.0 ml RIPAbuffer. Suspend the final pellet in 40 ul 2X SDS loading buffer,heated at 100'C for 5 minutes and recentifuge for 5 minutes in a microcentrifuge. Load 40uls of the supernatent onto a 8-16% gradient SDA-PAGE minigel. After electrophoresis, transfer the proteins to a nitrocellulose filter and block overnight in blocking buffer (3% BSA, 10 mM Tris pH 7.5, 100 mM NaC1, & 0.1% Tween-20). React the transferred proteins with Endothelin A & B receptor antibodies diluted 1:400 in blocking buffer for 1 hour at 25'C. Then add rabbit anti-sheep IgG HRP (KPL) diluted 1:5000 in blocking buffer for 1 hour at 25'C. Wash the nitrocellulose filter extensively. Detect the Endothelin A & B proteins using the Enhanced Chemiluminescent detection system by Amersham (tm).
Saponin Method:
Dulbecco's PBS (without Mg2+ or CA2+)
1% heat inactivated FCS (or RIA grade BSA)
0.1% w/v sodium azide
0.1% w/v saponin (Sigma cat#S-7900)
Adjust pH buffer to 7.4-7.6 and filter
see ref:
1)Sander, B., J. Andersson and U. Andersson, 1991:Assessment of cytokines by immunofluorsecence and the paraformaldehyde-saponin procedure:Immunol Rev 119:65-93
2) Anderson, U. and J. Andersson 1994. Immunolabelling of cytokine producing cells in tissues and suspension. In Cytokine Producing Cells eds. D. Fradelizie and D. Emelie. INSERM, Paris.32-49.
-immunogen sequence: AC-s-s-h-v-e-d-f-t-p-f-p-g-t-e-f-c-amide
-antibody purified by Protein G aff chromatography
-use: western blot, Eia
-immunogen: endothelin B-n-terminal receptor-GST fusion protein
sequence: G S N S S L M R F R T A E V
-antibody purified by Protein G affinity chromatography
-suitable for EIA, western blot (also humna/mouse/rat reactive)
-immunogen: Endothelin B receptor peptide-KLH conjugate
sequence: Ac-g-s-n-s-s-l-m-r-f-r-t-a-e-v-c-amide
-antibody purified by Protein G chromatography
-use: western blot, Eia
RDI Divison of researchd Industries Intl
San Jose, 95123 CA Snell ave 658
USA
or 408-780-0908
EMAIL:margaret@cellular-research.com
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