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Purified Mouse Anti-GRIP
Purified Mouse Anti-GRIP

Western blot analysis of GRIP expression on a rat cerebrum lysate (left panel). Rat Cerebrum Lysate (Cat. No. 611463) was stained with Purified Mouse Anti-GRIP (Cat. No. 611318/611319) at dilutions of 1: 1:1000, 1:2000, and 1:4000 (lanes 1, 2, and 3 respectively). GRIP expression was visualized with HRP Goat Anti-Mouse Ig (Cat. No. 554002) and appears as a 130 kDa band.

Purified Mouse Anti-GRIP

Immunofluorescence analysis of GRIP expression on undifferentiated SH-SY5Y cells (Human neuroblastoma; ATCC CRL-2266) (left) and differentiated SH-SY5Y cells (right).  Undifferentiated cells were seeded in a collagen coated 384-well imaging plate at ~ 8,000 cells per well.  After overnight incubation, cells were stained using the methanol fix/perm protocol (see Recommended Assay Procedure) and the Purified Mouse Anti-GRIP.  Differentiated cells were seeded in a 96-well, collagen coated imaging plate at ~ 5,000 cells per well.  Cells were incubated with 50 mM ATRA (Sigma-Aldrich, Cat.No. R2625) for 5 days, followed by 50 ng/ml BDNF (Sigma-Aldrich, Cat.No. B3795) for 5 days.  Differentiated cells were fixed and stained using the methanol fix/perm protocol, and Purified Mouse Anti-GRIP.  The second step reagent in both cases was Alexa Fluor® 488 goat anti-mouse Ig (Invitrogen).  The images were taken on a BD Pathway™ 855 or 435 Bioimager using a 20x objective.  This antibody also stained undifferentiated SK-N-SH (Human neuroblastoma; ATCC HTB-11), C6 (Rat glioma; ATCC CCL-107), U-87 MG (Human glioblastoma cells; ATCC HTB-14) and U-373 cells (Human glioblastoma cells; ATCC HTB-17; discontinued) using both the Triton-X 100 and methanol fix/perm protocols (see Recommended Assay Procedure).

Western blot analysis of GRIP expression on a rat cerebrum lysate (left panel). Rat Cerebrum Lysate (Cat. No. 611463) was stained with Purified Mouse Anti-GRIP (Cat. No. 611318/611319) at dilutions of 1: 1:1000, 1:2000, and 1:4000 (lanes 1, 2, and 3 respectively). GRIP expression was visualized with HRP Goat Anti-Mouse Ig (Cat. No. 554002) and appears as a 130 kDa band.

Immunofluorescence analysis of GRIP expression on undifferentiated SH-SY5Y cells (Human neuroblastoma; ATCC CRL-2266) (left) and differentiated SH-SY5Y cells (right).  Undifferentiated cells were seeded in a collagen coated 384-well imaging plate at ~ 8,000 cells per well.  After overnight incubation, cells were stained using the methanol fix/perm protocol (see Recommended Assay Procedure) and the Purified Mouse Anti-GRIP.  Differentiated cells were seeded in a 96-well, collagen coated imaging plate at ~ 5,000 cells per well.  Cells were incubated with 50 mM ATRA (Sigma-Aldrich, Cat.No. R2625) for 5 days, followed by 50 ng/ml BDNF (Sigma-Aldrich, Cat.No. B3795) for 5 days.  Differentiated cells were fixed and stained using the methanol fix/perm protocol, and Purified Mouse Anti-GRIP.  The second step reagent in both cases was Alexa Fluor® 488 goat anti-mouse Ig (Invitrogen).  The images were taken on a BD Pathway™ 855 or 435 Bioimager using a 20x objective.  This antibody also stained undifferentiated SK-N-SH (Human neuroblastoma; ATCC HTB-11), C6 (Rat glioma; ATCC CCL-107), U-87 MG (Human glioblastoma cells; ATCC HTB-14) and U-373 cells (Human glioblastoma cells; ATCC HTB-17; discontinued) using both the Triton-X 100 and methanol fix/perm protocols (see Recommended Assay Procedure).

Product Details
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BD Transduction Laboratories™
Glutamate Receptor Interacting Protein
Rat (QC Testing)
Mouse IgG1
Rat GRIP aa. 877-1067
Bioimaging, Western blot (Routinely Tested), Immunofluorescence (Tested During Development)
130 kDa
250 µg/ml
AB_398845
Aqueous buffered solution containing BSA, glycerol, and ≤0.09% sodium azide.
RUO


Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. Store undiluted at -20°C.

Recommended Assay Procedures

Bioimaging:

Methanol Procedure for a 96 well plate:

Remove media from wells. Add 100 µl/well fresh 3.7% Formaldehyde in PBS. Incubate for 10 minutes at room temperature (RT). Flick out and add 100 µl/well 90% methanol. Incubate for 5 minutes at RT. Flick out and wash twice with PBS. Flick out PBS and add 100 µl/well blocking buffer (3% FBS in PBS). Incubate for 30 minutes at RT. Flick out and add diluted antibody (diluted in blocking buffer). Incubate for 1 hour at RT. Wash three times with PBS. Flick out PBS and add second step reagent. Incubate for 1 hour at RT. Wash three times with PBS. Image sample.

Triton-X 100 Procedure for a 96 well plate:

Remove media from wells. Add 100 µl/well fresh 3.7% Formaldehyde in PBS. Incubate for 10 minutes at room temperature (RT). Flick out and add 100 µl/well 0.1% Triton-X 100. Incubate for 5 minutes at RT. Flick out and wash twice with PBS. Flick out PBS and add 100 µl/well blocking buffer (3% FBS in PBS). Incubate for 30 minutes at RT. Flick out and add diluted antibody (diluted in blocking buffer). Incubate for 1 hour at RT. Flick out and wash three times with PBS. Flick out and add second step reagent. Incubate for 1 hour at RT. Flick out and wash three times with PBS. Image sample.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
  3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  4. Sodium azide is a reversible inhibitor of oxidative metabolism; therefore, antibody preparations containing this preservative agent must not be used in cell cultures nor injected into animals. Sodium azide may be removed by washing stained cells or plate-bound antibody or dialyzing soluble antibody in sodium azide-free buffer. Since endotoxin may also affect the results of functional studies, we recommend the NA/LE (No Azide/Low Endotoxin) antibody format, if available, for in vitro and in vivo use.
  5. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  6. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
  7. Triton is a trademark of the Dow Chemical Company.
  8. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
611318 Rev. 2
Antibody Details
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32/GRIP

Rapid neuronal excitation within the CNS is mediated by the interactions of  receptors with their respective neurotransmitters, such as glutamate. Glutamate has a diverse array of receptors that are categorized into two distinct groups: ionotropic and metabotropic. Ionotropic receptors are ligand-gated channels and  can be subdivided into two classes: NMDA and AMPA receptors. AMPA receptors are composed of four homologous subunits (GluR1-4) that differentially combine to form a variety of receptor subtypes. Interactions of the subunits with cytoplasmic proteins mediate the transmission of extracellular signals. GRIP  (Glutamate Receptor Interacting Protein) specifically interacts with the C-terminus of the GluR2 subunit. GRIP lacks a catalytic domain, but contains seven PDZ domains which are motifs that mediate protein-protein interactions. Since only the fourth and fifth PDZ domains are utilized in interaction with GluR2, it is thought that the remaining PDZ domains interact with other unidentified proteins. Therefore, GRIP functions as an adaptor that links AMPA receptors to  cytoskeletal and/or signaling molecules and, in turn, clusters them at the synapse.

611318 Rev. 2
Format Details
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Purified
Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
Purified
611318 Rev.2
Citations & References
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Development References (4)

  1. Bowery NG, Brown DA. The cloning of GABA(B) receptors. Nature. 1997; 386(6622):223-224. (Biology). View Reference
  2. Dong H, O'Brien RJ, Fung ET, Lanahan AA, Worley PF, Huganir RL. GRIP: a synaptic PDZ domain-containing protein that interacts with AMPA receptors. Nature. 1997; 386(6622):279-284. (Biology). View Reference
  3. Fallon L, Moreau F, Croft BG, Labib N, Gu WJ, Fon EA. Parkin and CASK/LIN-2 associate via a PDZ-mediated interaction and are co-localized in lipid rafts and postsynaptic densities in brain. J Biol Chem. 2002; 277(1):486-491. (Biology: Western blot). View Reference
  4. Setou M, Seog DH, Tanaka Y. Glutamate-receptor-interacting protein GRIP1 directly steers kinesin to dendrites. Nature. 2002; 417(6884):83-87. (Biology: Immunofluorescence, Immunoprecipitation, Western blot). View Reference
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611318 Rev. 2

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Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

Non-IVD products are For Research Use Only. Not for use in diagnostic or therapeutic procedures.

 

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