APP antibody

Catalog No. ABIN95037

Product Details anti-APP Antibody

Target: APP (Amyloid beta (A4) Precursor Protein (APP))

Reactivity: Human, Mouse

Host: Rabbit

Clonality: Polyclonal

Conjugate: This APP antibody is un-conjugated

Application: Western Blotting (WB), ELISA, Immunohistochemistry (IHC), Immunofluorescence (IF), Fluorescence Microscopy (FM)

Purpose: Beta Amyloid Antibody

Cross-Reactivity (Details): This affinity purified antibody is directed against extracellular region of beta amyloid and is useful in determining its presence in various assays.

Characteristics: Synonyms: rabbit anti-Beta Amyloid Antibody, ß-amyloid, Amyloid beta A4 protein, Alzheimer disease amyloid protein, Beta amyloid, A-beta, ABPP, APPI, Beta-amyloid precursor protein

Purification: affinity purified antibody

Sterility: Sterile filtered

Immunogen:

Immunogen: This antibody was affinity purified from whole rabbit serum prepared by repeated immunizations with a synthetic peptide corresponding to an extracellular region of human beta amyloid conjugated to KLH using maleimide.

Immunogen Type: Conjugated Peptide

Isotype: IgG

Application Details

Application Notes:

Immunohistochemistry Dilution: 1:50-1:200

Application Note: Affinity purified anti-beta amyloid has been tested by ELISA, IHC, WB, and IF. A 45.8 kDa band is detected in western blot using whole tissue extracts and lysates from mouse and human.  In general, we recommend the use of 4 % PFA or 10 % formalin for fixation of tissues with IHC-paraffin or IHC-frozen application of this antibody.

Western Blot Dilution: 1:1,000-1:5000

ELISA Dilution: 1:10,000 - 1:30,000

IF Microscopy Dilution: 1:50-1:200

Other: User Optimized

Restrictions: For Research Use only

Independent Validation (UP)

Validation #104082 (Unfolding Profile)

by: NanoTemper Technologies

No.: #104082

Date: 07/23/2019

Antigen: ABETA

Method validated: Unfolding Profile

Positive Control: ABIN1043866

Notes:

Passed. ABIN1043866 showed T_i at 83.8°C and a clear unfolding profile with one unfolding event. This suggests that the antibody is properly folded and functional.

Validation Images

Unfolding profile of ABIN1043866. The fluorescence signal is plotted against temperature. The vertical line indicates the Ti at 83.8 °C.

Full Methods

Full Protocol:

• Load the undiluted sample into Tycho capillary (NanoTemper Technologies, TY-C001).
• Run Tycho measurement.

Experimental Notes:

Tycho is designed to run quick and precise protein quality check experiments. Tycho uses intrinsic protein fluorescence to follow protein unfolding while running a fast thermal ramp, yielding results in 3min. A protein’s unfolding behavior is characterized by various parameters, most notably the inflection temperature (T_i). The T_i can be used to identify properly folded protein, to compare different batches, or to analyze the influence of storage/transport conditions on a protein. An absence of T_i would suggest that the protein is already unfolded and therefore most likely nonfunctional.

Independent Validation (IHC)

Validation #104424 (Immunohistochemistry)

by: Prof. Merighi, Laboratory of Neurobiology, Department of Veterinary Sciences, University of Turin

No.: #104424

Date: 05/12/2022

Antigen: beta Amyloid

Lot Number: 42581

Method validated: Immunohistochemistry

Positive Control:

Adult (24 months) mouse brain

3-month-old Taconic mouse Model 1349 brain fixed in 4% paraformaldehyde

Negative Control:

We incubated slices overnight with the blocking solution only and then processed them with the secondary antibody.

Notes:

The beta Amyloid antibody (Amyloid beta) (N-Term) ABIN95037 works in IHC-P, especially at higher concentrations (1:20 and 1:50), and without the use of any antigen retrieval treatment.

Validation Images

Staining of beta-amyloid positive cells in the adult mouse brain using ABIN95037 at 40x magnification.

Full Methods

Primary Antibody: ABIN95037

Secondary Antibody: goat anti-rabbit AF488-conjugated antibody (Invitrogen by Thermo Fisher Scientific, A11034, lot 1971418)

Full Protocol:

• Perfuse mice with paraformaldehyde 4% in 0.1 M phosphate buffer pH 7.4 and post-fix in the same fixative for an additional 2 h at RT.
• Wash, dehydrate, and embed samples in paraffin wax.
• Wash several times with 0.01 M PBS.
• Cut intestines and brain with a microtome into 6µm sections and mount on glass slides.
• After paraffin removal, incubate sections for 1 h at RT in PBS containing 1% albumin from chicken egg white (Sigma, A5378) and 0.3% Triton-X-100 (BioRad, 161-0407, lot 00583) to block non-specific binding sites.
• Incubate sections with primary anti-beta amyloid (N-Term) (antibodies Online, ABIN95037, lot 42581) diluted 1:20, 1:50, 1:100, and 1:200 in 0.1 M PBS-BSA-PLL ON at RT.
• Wash sections 3x 5 min with 0.01 M PBS.
• Incubate sections with secondary goat anti-rabbit AF488-conjugated antibody (Invitrogen by Thermo Fisher Scientific, A11034, lot 1971418) diluted 1:500 in 0.1 M PBS for 1 h at RT.
• Wash sections 3x 5 min with 0.01M PBS.
• Mount specimens in Fluoroshield (Sigma, F6182, lot MKCB0153V).
• Acquire images with a fluorescence microscope and appropriate filter settings for AF488, e.g. Leica DM 6000B fluorescence microscope equipped with a digital camera at 40x magnification.

Experimental Notes:

Antigen retrieval treatment was also tested. In this case, sections were processed for microwave antigen retrieval for 10 min (95-100 °C) in 10 mM sodium citrate buffer (pH 6.0). After 20 min of spontaneous cooling, sections were washed twice for 5 min with distilled water and for 5 min with PBS.

Handling

Format: Liquid

Buffer:

Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2

Stabilizer: None

Preservative: 0.01 % (w/v) Sodium Azide

Preservative: Sodium azide

Precaution of Use: This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Storage: 4 °C,-20 °C

Storage Comment: Store vial at -20° C prior to opening. Aliquot contents and freeze at -20° C or below for extended storage. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4° C as an undiluted liquid. Dilute only prior to immediate use.

Expiry Date: 12 months

References for anti-APP antibody (ABIN95037)

Frenkel, Balass, Solomon: "N-terminal EFRH sequence of Alzheimer's beta-amyloid peptide represents the epitope of its anti-aggregating antibodies." in: Journal of neuroimmunology, Vol. 88, Issue 1-2, pp. 85-90, (1998) (PubMed).

Target Details for APP

Target: APP (Amyloid beta (A4) Precursor Protein (APP))

Alternative Name: APP (APP Products)

Background: Background: Beta amyloid, often abbreviated as A-beta, is a protein that builds up in the brains of persons with Alzheimer's disease, collecting in clumps called plaques or senile plaques. While some researchers question whether beta amyloid is the cause of the dementia, most agree that it is involved in the disruption of thinking that is a hallmark of the disease. In some cases of familial Alzheimer's disease, mutations in genes for the proteins called the presenilins lead to increased production of amyloid. Researchers have been looking at how presenilin-1 in particular contributes to the excess buildup of beta amyloid. Presenilin-1 apparently acts to increase the activity of gamma-secretase, an enzyme that changes a normal protein (amyloid precursor protein or APP) into beta amyloid itself. Furthermore, presenilin-1 might be gamma-secretase. Anti-Beta Amyloid Antibody is useful for researchers interested in TLR signaling and Alzheimer's research.

Gene ID: 351

NCBI Accession: NP_000475

UniProt: P05067

Pathways: Caspase Cascade in Apoptosis, EGFR Signaling Pathway, Transition Metal Ion Homeostasis, Skeletal Muscle Fiber Development, Toll-Like Receptors Cascades, Feeding Behaviour