ELISA Kit was purchased from R&D Systems (Minneapolis, MN). Goat antihuman hScrib and mouse antihuman E-cadherin antibodies were purchased from R&D Systems (Minneapolis, MN). Donkey antigoat and rabbit antimouse secondary antibodies (Labelled Polymer-HRP, Dako). Allexa 488 and Alexa-568 conjugated antibodies (Invitrogen, Eugene, OR).
Polymer hrp
Manufactured by Agilent Technologies
Sourced in United States
Polymer-HRP is a horseradish peroxidase (HRP) conjugate designed for use in various immunoassay applications. It functions as a label or detection reagent to facilitate the visualization and quantification of target analytes in such assays.
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Lab products found in correlation
3 protocols using polymer hrp
1
ELISA and Immunofluorescence Assay
2
Double Immunofluorescence for Bcl-6 and CD3/CD20
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Double immunofluorescence (IF) staining for Bcl-6 in combination with CD3 (clone LN10, mmAb, Novocastra) or CD20 was carried out with sequential immunostaining, as described previously [32 (link)]. Sections were prepared and antigens were unmasked as detailed above. After 1-hour treatment with anti-Bcl-6 primary Ab, the slides were incubated using anti-mouse IgG(Fab)2 as secondary Ab coupled to polymer-HRP (Dako), followed by a tetramethylrhodamine- (TMR-) conjugated tyramide reagent of the fluorescent amplification kit (TSR-TMR System, Perkin Elmer Life Science, Boston, MA, USA) to visualize the red nuclear fluorescence. The second layer of the double IF staining was applied with anti-CD3 or anti-CD20 primary Abs plus biotinylated anti-mouse secondary IgG F(ab')2 followed by streptavidin-fluorescein isothiocyanate (FITC). Nuclear counterstaining was made with DAPI (blue fluorescence, Vector Laboratories). Images were obtained using a Zeiss AXIO Imager Z2 microscope (Carl Zeiss Microscopy GmbH, Jena, Germany) equipped with the following objectives: 10x/0.3 NA; 20x/0.5 NA. For transferring and editing images, Isis software (MetaSystems Group Inc., Newton, MA, USA) and Adobe Photoshop CS5 version 12.0 were used.
3
Neurocan Expression in Murine Embryos
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A collection of brain tissues from day 14 embryos and newborn mice was employed to evaluate neurocan levels. The slides were processed for indirect peroxidase immunohistochemistry in the following way: the sections were deparaffinized and rehydrated, and then rinsed in phosphate-buffered saline (PBS) containing 1% Tween-20. For neurocan detection (anti-neurocan antibody from R&D Systems, Ref. AF5800), the sections were heated in high pH Envision FLEX target retrieval solution at 80 °C for 20 min and incubated for 20 min at room temperature in the same solution. Endogenous peroxidase activity (3% H 2 O 2 ) and non-specific binding (33% fetal calf serum) were blocked and the sections were incubated overnight at 4 °C with a primary antibody (1:100 dilution) as described above. A secondary antibody labeled with polymer-HRP (DAKO) was used. 3, 3′-Diaminobenzidine was employed as a chromogen. Selected slides were counterstained with hematoxylin.
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