NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

Novus Biologicals | Catalog # NB110-81529

Novus Biologicals
100% Guarantee

Key Product Details

Species Reactivity

Validated:

Human, Mouse, Rat, Porcine, Opossum, Primate, Rabbit

Cited:

Human, Mouse, Rat, Porcine, Opossum, Primate

Applications

Validated:

Knockout Validated, Immunohistochemistry, Western Blot, ELISA, Flow Cytometry, Immunocytochemistry/ Immunofluorescence, Simple Western, Immunoprecipitation, Knockdown Validated

Cited:

Knockout Validated, Western Blot, ELISA, Flow Cytometry, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, IF/IHC, Knockdown Validated, SDS-Page

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG2A Clone # 14D5

Format

BSA Free
View all NHE3/SLC9A3 Primary Antibodies »

Product Specifications

Immunogen

Synthetic peptide made to a region surrounding the Serine 552 portion of rat NHE3. [UniProt# P26433]

Reactivity Notes

Porcine reactivity reported in scientific literature (PMID: 29510506).

Modification

p Ser552

Specificity

This antibody detects NHE3 posphorylated at Serine 552, and does not cross-react with the non-phosphopeptide.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG2A

Theoretical MW

85 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

Western Blot: NHE3/SLC9A3 [p Ser552] Antibody (14D5) [NB110-81529] - Detection of NHE3 [p Ser552] in human kidney lysate.
Immunohistochemistry: NHE3/SLC9A3 [p Ser552] Antibody (14D5) [NB110-81529] - Mouse intestine, image courtesy of verified customer review.
Simple Western: NHE3/SLC9A3 [p Ser552] Antibody (14D5) [NB110-81529] - Simple Western lane view shows a specific band for NHE3 in 0.5 mg/ml of Hek293 lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.

Applications for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

Application
Recommended Usage

ELISA

1:100-1:2000

Flow Cytometry

reported in scientific literature (PMID 21255452)

Immunocytochemistry/ Immunofluorescence

1:50-1:1000

Immunohistochemistry

1:50-1:1000

Immunoprecipitation

1:10-1:500

Knockdown Validated

reported in scientific literature (PMID 31276916)

Knockout Validated

reported in scientific literature (PMID 31276916)

Simple Western

1:1000

Western Blot

1-5 ug/ml
Application Notes
In Western Blot, a band can be seen at approx. 85 kDa.

In Simple Western only 10 - 15 uL of the recommended dilution is used per data point.
See Simple Western Antibody Database for Simple Western validation: Tested in Hek293 lysate 0.5 mg/mL, separated by Size, antibody dilution of 1:1000, apparent MW was 81 kDa. Separated by Size-Wes, Sally Sue/Peggy Sue.

Reviewed Applications

Read 1 review rated 4 using NB110-81529 in the following applications:

Flow Cytometry Panel Builder

Bio-Techne Knows Flow Cytometry

Save time and reduce costly mistakes by quickly finding compatible reagents using the Panel Builder Tool.

Advanced Features

  • Spectra Viewer - Custom analysis of spectra from multiple fluorochromes
  • Spillover Popups - Visualize the spectra of individual fluorochromes
  • Antigen Density Selector - Match fluorochrome brightness with antigen density
Build Your Panel Now
Flow Cytometry

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

Tris-Glycine and 0.15M NaCl

Format

BSA Free

Preservative

0.05% Sodium Azide

Concentration

2 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: NHE3/SLC9A3

Na+/H+ exchanger (NHE) family members are integral membrane proteins which play a key role in catalyzing the extrusion of intracellular proton (H+) ions in exchange for extracellular sodium (Na+) ions for regulation of cellular pH as well as transepithelial ion/water transport. NHE3 is one of the best characterized NHEs that is a major proton extruding system driven by inward Na+ ion chemical gradient and is implicated in pH regulation to eliminate acids generated during metabolism or to tackle any potential adverse conditions in tissue microenvironment. NHE3 exists as a multi-pass membrane protein in cell membranes and is expressed most abundantly in colon and the small intestine, followed by the kidneys and the stomach, as well as in ventrolateral region of the medulla oblongata. In intestinal epithelial cells, it localizes to the ileal brush border and SGK1 mediated phosphorylation at Ser-661 has been shown to be associated with its enhanced cell membrane localization. NHE3 binds to SLC9A3R1 and SLC9A3R2, and interacts with CHP1, CHP2, SHANK2 PDZD3 and (interactions may decrease in response to elevated calcium ion levels). NHE3 is regulated by several hormones, neurotransmitters, and associated signaling systems such as cAMP, cGMP, and elevated intracellular calcium via mechanisms that are largely partially understood. NHE3 knock-out mice suffers from chronic diarrhea and altered salt as well as water homeostasis.

Long Name

Sodium/Hydrogen Exchanger 3

Alternate Names

SLC9A3

Gene Symbol

SLC9A3

Additional NHE3/SLC9A3 Products

Product Documents for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

Download SDS

Product Specific Notices for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Related Research Areas

Ion Channels and Regulators

Citations for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

Customer Reviews for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free (1)

4 out of 5
1 Customer Rating
5 Stars
0%
4 Stars
100%
3 Stars
0%
2 Stars
0%
1 Stars
0%

Have you used NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free?

Submit a review and receive an Amazon gift card!

$25/€18/£15/$25CAN/¥2500 Yen for a review with an image

$10/€7/£6/$10CAN/¥1110 Yen for a review without an image

Submit a review
Amazon Gift Card

Customer Images

Showing  1 - 1 of 1 review Showing All
Filter By:
Clear Filters X
  • NHE3/SLC9A3 [p Ser552] Antibody (14D5)
    Name: Anjum Jafri
    Application: Immunohistochemistry-Paraffin
    Sample Tested: Mouse Intestine
    Species: Mouse
    Verified Customer | Posted 03/16/2017
    NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free NB110-81529

There are no reviews that match your criteria.

Showing  1 - 1 of 1 review Showing All

Protocols

View specific protocols for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free (NB110-81529):


Western Blot Protocol

1. Perform SDS-PAGE (4-12% MOPS) on samples to be analyzed, loading 30 ug of total protein per lane.
2. Transfer proteins to Nitrocellulose according to the instructions provided by the manufacturer of the transfer
apparatus.
3. Rinse membrane with dH2O and then stain the blot using Ponceau S for 1-2 minutes to access the transfer of proteins onto the nitrocellulose membrane. Rinse the blot in water to remove excess stain and mark the lane locations and locations of molecular weight markers using a pencil.
4. Rinse the blot in TBS for approximately 5 minutes.
5. Block the membrane using 5% NFDM + 1% BSA in TBS + Tween, 1 hour at RT.
6. Rinse the membrane in dH2O and then wash the membrane in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each.
7. Dilute the mouse anti-NHE3 primary antibody (NB 110-81529) in blocking buffer and incubate 1 hour at room temperature.
8. Rinse the membrane in dH2O and then wash the membrane in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each.
9. Apply the diluted mouse-IgG HRP-conjugated secondary antibody in blocking buffer (as per manufacturers
instructions) and incubate 1 hour at room temperature.
10. Wash the blot in wash buffer [TBS + 0.1% Tween] 3 times for 10 minutes each (this step can be repeated as required to reduce background).
11. Apply the detection reagent of choice in accordance with the manufacturers instructions (Pierce ECL).

Note: Tween-20 can be added to the blocking or antibody dilution buffer at a final concentration of 0.05-0.2%, provided it does not interfere with antibody-antigen binding.

Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.

FAQs for NHE3/SLC9A3 [p Ser552] Antibody (14D5) - BSA Free

Showing  1 - 2 of 2 FAQs Showing All

  • Q: The molecular weight of NHE3 is 92kDa but I see a band above 100 kDa.  I am confused, because the size of these protein should not be greater than 100KD. Can you please help explain?  

    A: Higher than predicted molecular weight seen on WB is not uncommon if the protein is posttranslationally modified, especially glycosylated, and such is the case for NHE3. Please note that NHE3 gets phosphorylated and glycosylated (Uniprot ID P48764) and potentially, these modifications are adding ~10kDa extra to the predicted molecular weight. If you want to confirm it 100%, you could try de-glycosylating your protein and then performing a WB on it. 

  • Q: What application was used to test on mouse species? Was it WB, ELISA, IHC-paraffin, IHC-frozen, ICC/IF, or etc?

    A: We tested each lot in Western blot on human, mouse, and rat colon, small intestine and kidney samples. There were bands expressed around 25kDa and 50kDa, with weaker bands at approx. 80-90 kDa observed. This antibody is guaranteed to work in samples from mouse that are expected to express the protein. Since it is a phospho specific antibody the customer must use BSA to block and not milk.

  • Q: The molecular weight of NHE3 is 92kDa but I see a band above 100 kDa.  I am confused, because the size of these protein should not be greater than 100KD. Can you please help explain?  

    A: Higher than predicted molecular weight seen on WB is not uncommon if the protein is posttranslationally modified, especially glycosylated, and such is the case for NHE3. Please note that NHE3 gets phosphorylated and glycosylated (Uniprot ID P48764) and potentially, these modifications are adding ~10kDa extra to the predicted molecular weight. If you want to confirm it 100%, you could try de-glycosylating your protein and then performing a WB on it. 

  • Q: What application was used to test on mouse species? Was it WB, ELISA, IHC-paraffin, IHC-frozen, ICC/IF, or etc?

    A: We tested each lot in Western blot on human, mouse, and rat colon, small intestine and kidney samples. There were bands expressed around 25kDa and 50kDa, with weaker bands at approx. 80-90 kDa observed. This antibody is guaranteed to work in samples from mouse that are expected to express the protein. Since it is a phospho specific antibody the customer must use BSA to block and not milk.

Showing  1 - 2 of 2 FAQs Showing All
View all FAQs for Antibodies