Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed)

Novus Biologicals | Catalog # NBP1-72944

Novus Biologicals

Key Product Details

Species Reactivity

Rabbit

Applications

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Fluorophore-linked immunosorbent assay, Immunocytochemistry/ Immunofluorescence

Label

DyLight 488 (Excitation = 493 nm, Emission = 518 nm)

Antibody Source

Polyclonal Goat IgG

Format

Pre-adsorbed
View all IgG (H+L) Secondary Antibodies »

Product Specifications

Immunogen

Rabbit IgG whole molecule

Specificity

This antibody was pre-adsorbed against Bovine, Chicken, Goat, Guinea Pig, Hamster, Horse, Human, Mouse, Rat, and Sheep Serum Proteins. No reaction was observed against Bovine, Chicken, Goat, Guinea Pig, Hamster, Horse, Human, Mouse, Rat and Sheep Serum Proteins. This antibody will react with heavy chains of rabbit IgG and with light chains of most rabbit immunoglobulins.

Clonality

Polyclonal

Host

Goat

Isotype

IgG

Description

Store vial at 4C prior to restoration. For extended storage aliquot contents and freeze at -20C or below. 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 4C as an undiluted liquid. Dilute only prior to immediate use.

This product was prepared from monospecific antiserum by immunoaffinity chromatography using Rabbit IgG coupled to agarose beads followed by solid phase adsorption(s) to remove any unwanted reactivities. Assay by immunoelectrophoresis resulted in a single precipitin arc against anti-Goat Serum, Rabbit IgG and Rabbit Serum

Scientific Data Images

Western Blot: Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed) [NBP1-72944] - Western Blot of Unconjugated Goat anti-Rabbit IgG (H+L) Secondary antibody [DyLight 488] (Pre-adsorbed). Lane M: 3 ul Molecular Ladder. Lane 1: Rabbit IgG whole molecule. Lane 2: Rabbit IgG F(ab) Fragment. Lane 3: Rabbit IgG F(c) Fragment. Lane 4: Rabbit IgM Whole Molecule. Lane 5: Normal Rabbit Serum. All samples were reduced. Load: 50 ng per lane. Block for 30 min at RT. Primary Antibody: Goat anti-Rabbit IgG (H+L) Secondary antibody [DyLight 488] (Pre-adsorbed) 1:1,000 for 60 min at RT. Secondary antibody: Anti-Goat IgG (DONKEY) Peroxidase Conjugated Antibody 1:40,000 in blocking buffer for 30 min at RT. Predicted/Observed Size: 25 and 50 kDa for Rabbit IgG and Serum, 25 kDa for F(c) and F(ab), 70 and 23 kDa for IgM. Rabbit F(c) migrates slightly higher.
Immunocytochemistry/Immunofluorescence: Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed) [NBP1-72944] - Myosin expression in HeLa cells. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature. The secondary antibody was diluted in 1% BSA in PBS at 1:500 and incubated for 20 minutes at room temperature. Image from verified customer review.
Immunocytochemistry/Immunofluorescence: Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed) [NBP1-72944]
Immunocytochemistry/Immunofluorescence: Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed) [NBP1-72944] - DyLight 488 conjugated anti-Rabbit IgG was used to demonstrate 2 color STED immunofluorescence microscopy. Methanol fixed A431 cells were blocked with normal goat serum. The cells were then probed with 0.4 ug/mL final concentration of anti-HDAC and detected with 0.2 ug/mL DyLight488 conjugated Anti-RABBIT IgG [GOAT] secondary antibody (colored GREEN). Also shown in this 2-color STED image is a-tubulin monoclonal antibody [MOUSE] detected with ATTO 425 conjugated anti-MOUSE IgG [GOAT] secondary antibody (colored RED).

Applications

Application
Recommended Usage

Fluorophore-linked immunosorbent assay

1:20000

Immunocytochemistry/ Immunofluorescence

1:5000

Immunohistochemistry

1:50 - 1:200

Immunohistochemistry-Paraffin

1:20 - 1:50

Western Blot

1:10000-1:25000
Application Notes
This secondary antibody is designed for immunofluorescence microscopy, fluorescence based plate assays (FLISA) and fluorescent western blotting. This product is also suitable for multiplex analysis, including multicolor imaging, utilizing various commercial platforms. The emission spectra for this DyLight(TM) conjugate match the principle output wavelengths of most common fluorescence instrumentation.

Reviewed Applications

Read 1 review rated 5 using NBP1-72944 in the following applications:

Formulation, Preparation, and Storage

Purification

Multi-step

Reconstitution

Reconstitute with 100 ul deionized water (or equivalent).

Formulation

Lyophilized from 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2, 10 mg/mL Bovine Serum Albumin (BSA) - Immunoglobulin and Protease free

Format

Pre-adsorbed

Preservative

0.01% Sodium Azide

Concentration

LYOPH mg/ml

Shipping

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

Stability & Storage

Store lyophilized antibody at 4C in the dark. Aliquot reconstituted liquid and store at -20C. Avoid freeze-thaw cycles.

Calculators

The reconstitution calculator allows you to quickly calculate the volume of a reagent to reconstitute your vial. Simply enter the mass of reagent and the target concentration and the calculator will determine the rest.

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Background: IgG (H+L)

Antibodies, also known as immunoglobulins (Igs) are critical for immunity and are grouped into five primary classes: IgG, IgM, IgA, IgD, and IgE. The most abundant antibody isotype is immunoglobulin G (IgG) with concentrations ranging from 7.5-22 mg/ml in human serum and has a molecular weight of 150 kDa. The major effector functions of IgG include neutralization, opsonization, complement fixation and antibody dependent cell-mediated cytotoxicity (ADCC). This monomeric immunoglobulin, expressed on the surface of mature B cells, is often depicted as a Y-shape and comprised of 2 heavy chains and 2 light chains linked by disulfide bonds. The heavy chain is type gamma including subtypes gamma 1, gamma 2, gamma 3, and gamma 4 while the light chain is either a kappa or lambda chain. An IgG molecule has two antigen binding sites, each consisting of a heavy and light chain N-terminal variable domain. When combined with the constant heavy chain 1 (Ch1) and the constant light chain domains, it forms the fragment antigen-binding (Fab) region (2 per antibody). The remaining domains (Ch2-Ch4) of both heavy chains make up the Fc region and contain a site for covalently linking an enzymatic or fluorochrome probe, such as HRP or Janelia Fluor 549, for target detection and visualization (1,2,3).

The 4 IgG subclasses, sharing 95% amino acid identity, include IgG1, IgG2, IgG3, and IgG4 for humans and IgG1, IgG2a, IgG2b, and IgG3 for mice. The relative abundance of each human subclass is 60% for IgG1, 32% for IgG2, 4% for IgG3, and 4% for IgG4. In an IgG deficiency, there may be a shortage of one or more subclasses (4).

References

1. Painter RH. (1998) Encyclopedia of Immunology (Second Edition). Elsevier. 1208-1211

2. Chapter 9 - Antibodies. (2012) Immunology for Pharmacy. Mosby 70-78

3. Schroeder H, Cavacini, L. (2010) Structure and Function of Immunoglobulins. J Allergy Clin Immunol. 125(2 0 2): S41-S52. PMID: 20176268

4. Vidarsson G, Dekkers G, Rispens T. (2014) IgG subclasses and allotypes: from structure to effector functions. Front Immunol. 5:520. PMID: 25368619

Additional IgG (H+L) Products

Product Documents

Certificate of Analysis

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

Product Specific Notices

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

Citations for Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed)

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  • Name: Anonymous
    Application: Immunofluorescence
    Sample Tested: hela
    Species: Human
    Verified Customer | Posted 11/12/2014
    myosin
    Goat anti-Rabbit IgG (H+L) Secondary Antibody [DyLight 488] (Pre-adsorbed) NBP1-72944

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