Rabbit Anti Human Betacellulin polyclonal, antigen affinity

///Rabbit Anti Human Betacellulin polyclonal, antigen affinity

Rabbit Anti Human Betacellulin polyclonal, antigen affinity

$150.00$190.00

SKU: RKAP016 Category: Tags: ,

accession P35070


Applications

ELISA:This antibody can be used at 2 μg/mL jointly with biotinylated Human Betacellulin antibody to detect Human Betacellulin. The detection limit for recombinant Human Betacellulin is approximately 0.2 ng/well.

Western Blot:This antibody can be used at 0.2 μg/mL with the appropriate secondary reagents to detect Human Betacellulin. The detection limit for recombinant Human Betacellulin is approximately 0.2 ng/lane.

Source This antibody was produced in rabbits immunized with recombinant Human Betacellulin.
Species reactivity Human
Purification The specific antibody was purified from Rabbit sera by using immobilized recombinant Human Betacellulin affinity chromatography.
Presentation Lyophilized from PBS, pH 7.2.
Storage The lyophilized antibody is stable for at least 1 year from date of receipt at -20° C. Upon reconstitution, this antibody can be stored in working aliquots at 2° - 8° C for one month, or at -20° C for 12 months without detectable loss of activity. Avoid repeated freeze/thaw cycles.
Usage This antibody product is for research purposes only.It may not be used for therapeutics or diagnostic purposes.


Interactor P00533 EGFR_HUMAN
Molecular function Growth-factor
Molecular function Mitogen

Methods

Western Blot Analysis

  • Total proteins were isolated from PDMSCs and chorionic villous explants using 1X Radio Immuno-precipitation Assay .
  • Fifty µg of total protein from PDMSCs and villous explants were processed by SDS-page electrophoresis on 4–12% polyacrylamide pre-cast gradient gels .
  • Next, proteins were transferred onfluoride (PVDF) membranes and probed at room temperature with primary antibodies using the SnapID system following manufacturer instructions.
  • Primary antibodies were: mouse monoclonal anti-human MIF (1∶1000 dilution& , , ), rabbit polyclonal anti-human VEGF (1∶1000 dilution& , , ), rabbit polyclonal anti-human soluble VEGFeceptor (sFlt-1, 1∶250 dilution , ), rabbit polyclonal anti-human Ki67 (1∶500 dilution , ), goat polyclonal anti-human Cytokeratin 7 (1∶500 dilution , ), mouse monoclonal anti-human C45 (1∶500 dilution , ) and mouse monoclonal anti-human C166 (1∶200 dilution , ).
  • Biotinylated secondary antibodies were goat anti-mouse for MIF, CD45 and CD166 (1∶1000 dilution , ) and donkey anti-rabbit for VEGF,…
  • Total proteins were isolated from PDMSCs and chorionic villous explants using 1X Radio Immuno-precipitation Assay .
  • Fifty µg of total protein from PDMSCs and villous explants were processed by SDS-page electrophoresis on 4–12% polyacrylamide pre-cast gradient gels .
  • Next, proteins were transferred onfluoride (PVDF) membranes and probed at room temperature with primary antibodies using the SnapID system following manufacturer instructions.
  • Primary antibodies were: mouse monoclonal anti-human MIF (1∶1000 dilution& , , ), rabbit polyclonal anti-human VEGF (1∶1000 dilution& , , ), rabbit polyclonal anti-human soluble VEGFeceptor (sFlt-1, 1∶250 dilution , ), rabbit polyclonal anti-human Ki67 (1∶500 dilution , ), goat polyclonal anti-human Cytokeratin 7 (1∶500 dilution , ), mouse monoclonal anti-human C45 (1∶500 dilution , ) and mouse monoclonal anti-human C166 (1∶200 dilution , ).
  • Biotinylated secondary antibodies were goat anti-mouse for MIF, CD45 and CD166 (1∶1000 dilution , ) and donkey anti-rabbit for VEGF, sFlt-1 and Ki67 (1∶1000 dilution , ) and donkey anti-goat for Cytokeratin 7 (1∶100 dilution , ).
  • Blots were visualized using the QDot reagent kit ( by , ) following manufacturer instructions.

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Sandwich ELISA

  • 96-well maxisorp plates were coated with the capture antibody overnight, then blocked with 5% FCS in PBS for 1 hour, then washed.
  • Then, the protein standard was added, and plates were incubated for 2 hrs at 37°C.
  • Plates were washed and incubated with a biotinylated detection antibody for 1 hour at 37°C.
  • After washing, plates were incubated with HRP-conjugated streptavidin for 30 min at room temperature and washed again.
  • Plates were developed using the tetramethylbenzidine peroxidase substrate system and absorbance was measured at 450-615 nm using an automated plate reader.

Western Blot

  • The Recombinant immunogen was migrated by 12% SDS-PAGE and transferred to a 0.2 um PVDF membrane.
  • After blocking the membrane in blocking buffer (5% milk powder in 20 mM Tris-HCl pH 7.5, 500 mM NaCl, 0.1% (v/v) Tween 20 , the membrane was incubated with primary antibody ( at 4°C overnight.
  • Peroxidase-linked secondary anti-rabbit were used to detect the bound primary antibodies.
  • Enhanced chemiluminescence (ECL) reagents were used to visualize