First Class Secondary Antibodies

Jackson ImmunoResearch

For over 37 years Jackson ImmunoResearch have specialised in the manufacture of secondary antibodies, conjugates and purified immunoglobulins. Their products are made by scientists for scientists and are considered by many to be the gold standard. With their state of the art production facility in rural Pennsylvania and there European hub in Cambridgeshire they serve Research Institutes and Universities throughout the world.

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Technical Centre

Download the JIR Catalogue, Product Handouts, White Papers and Posters

New! Anti-Camelid Secondary Antibodies

At Jackson ImmunoResearch, our mission is to develop the widest range of high quality secondary antibodies for the life science community. To support the growing adoption of camelid-derived heavy chain variable domain (VHH) technologies in a multitude of applications, including next-generation immunotherapeutics, medical imaging, analytical and diagnostic assay development and research discovery, JIR offers three new secondary antibody specificities for detection and quantification of camelid antibodies.

Immunoglobulins Poster

Download our guide to Immunoglobulins Structure and Function

Multiple Labelling Poster

Download our guide to Multiple Labeling with Secondary Antibodies

Flyers

Guides

See JIR Secondary antibodies in action

Contact marketing@stratech.co.uk for more information on any of the following images.

Anti-Alpaca IgG and VHH domain Secondary Antibodies

Find out about our Secondary Antibodies

Mouse on Mouse Labeling with Fab Fragment Blocking

Anti-Camelid Secondary Antibodies

Having manufactured and supplied secondary antibodies and related reagents for 34 years, Jackson ImmunoResearch know the best practices to employ, as well as the pitfalls to be avoided in their use. In these articles we offer the benefit of our experience with useful and practical advice.

Featured Products

Product Code Product Description
115-545-003 Alexa Fluor® 488-AffiniPure Goat Anti-Mouse IgG (H+L)
115-035-003 Peroxidase-AffiniPure Goat Anti-Mouse IgG (H+L)
115-545-166 Alexa Fluor® 488-AffiniPure Goat Anti-Mouse IgG (H+L) (min X Hu,Bov,Hrs,Rb,Rat Sr Prot)
715-545-151 Alexa Fluor® 488-AffiniPure Donkey Anti-Mouse IgG (H+L) (min X Bov,Ck,Gt,GP,Sy Hms,Hrs,Hu,Rb,Rat,Shp Sr Prot)
715-035-151 Peroxidase-AffiniPure Donkey Anti-Mouse IgG (H+L) (min X Bov,Ck,Gt,GP,Sy Hms,Hrs,Hu,Rb,Rat,Shp Sr Prot)
111-545-003 Alexa Fluor® 488-AffiniPure Goat Anti-Rabbit IgG (H+L)
111-035-003 Peroxidase-AffiniPure Goat Anti-Rabbit IgG (H+L)
111-545-144 Alexa Fluor® 488-AffiniPure Goat Anti-Rabbit IgG (H+L) (min X Hu,Ms,Rat Sr Prot)
111-035-144 Peroxidase-AffiniPure Goat Anti-Rabbit IgG (H+L) (min X Hu,Ms,Rat Sr Prot)
711-545-152 Alexa Fluor® 488-AffiniPure Donkey Anti-Rabbit IgG (H+L) (min X Bov,Ck,Gt,GP,Sy Hms,Hrs,Hu,Ms,Rat,Shp Sr Prot)
711-035-152 Peroxidase-AffiniPure Donkey Anti-Rabbit IgG (H+L) (min X Bov,Ck,Gt,GP,Sy Hms,Hrs,Hu,Ms,Rat,Shp Sr Prot)

FAQs

Manufacturing Questions

1. How do you determine protein concentration?

Jackson ImmunoResearch uses the Biuret assay to determine protein concentration.
If protein concentration is obtained by using the mass extinction coefficient of 1.4 for a 1 mg/ml solution at OD280, the result will be 10-15% lower than the concentration obtained by the Biuret assay.

2. What is the molecular weight of my product?

Jackson ImmunoResearch does not determine molecular weights by analytical methods. An approximate molecular weight of 160, 110 and 50 kDa can be used for whole IgG, F(ab’)2 and Fab fragment antibodies, respectively. To obtain an estimated molecular weight of any other product of interest, please contact Technical Service at help@jacksonimmuno.com.

3. Do you make custom antibodies?

For purchase of OEM products that have custom specifications (e.g. custom adsorptions, conjugations), please contact Technical Service at technical@stratech.co.uk. Jackson ImmunoResearch does not provide polyclonal or monoclonal antibody development or production services.

4. Are your products endotoxin-free?

Jackson ImmunoResearch does not assay products for endotoxin content. For applications requiring low endotoxin consider using an endotoxin removal column, available from Thermo Fisher, Charles River Laboratories, and other suppliers.

5. Why is there a discrepancy regarding the protein concentration, volume of dH2O used to reconstitute the lyophilized powder, and the nominal fill size reported on the spec sheet?

For products that are sold by weight (mg), Jackson ImmunoResearch fills vials with slightly more product that the nominal mg amount (“Size” on the spec sheet). To calculate the actual product amount in a vial, multiply the protein concentration indicated on the spec sheet by the volume of dH2O needed to rehydrate the lyophilized pellet.

General Application Questions

1. What are common causes of background?

Background may be caused by the primary antibody. Perform a control experiment excluding the primary antibody to isolate the secondary antibody as the cause of background.
Improper blocking of the tissues or cells.
Cross-reactivity of the labeled secondary antibodies with endogenous immunoglobulins on the tissues or cells.
Inadequate washing.
Reactivity of the labeled secondary antibody with immunoglobulins in the diluent.
Not diluting the secondary antibodies far enough.

2. What are some causes of weak signal?

There is insufficient antigen present and an amplification protocol may be needed.
Change from a more sensitive detection method to a less sensitive detection method.
The primary antibody is diluted too far.
Enzyme activity is inhibited.
The secondary antibody does not recognize certain primary antibodies well.
The secondary antibody cross-reacts with immunoglobulins in the diluent.

3. What diluent is recommended for the secondary antibody?

The antibodies can be diluted in buffers such as PBS or TBS. A detergent such as Tween 20 (0.05% v/v) can be included to reduce non-specific binding. We recommend making the working dilution fresh on the day of use. Carrier proteins such as BSA or normal serum are not necessary, and in some cases can increase background staining due to protein-protein interactions.
Read more about diluents in our guide to blocking, controls and diluents.

4. What blocking buffer is recommended?

For most applications, we recommend blocking with normal serum (5% v/v) from the same species as the host of the secondary antibody. Other commonly used reagents such as BSA and non-fat dry milk are suitable for some applications, but may increase background if the secondary antibody is directed against goat, sheep, or horse.
Read more about blocking in our guide to blocking, controls and diluents.

5. What incubation time is recommended for the secondary antibody?

Incubation times of 30-60 minutes are recommended for surface interactions such as ELISA or Western blotting. For staining tissue, incubation times should be increased as necessary to allow for tissue penetration. Total wash time should be approximately equal to incubation time to allow unreacted antibody to diffuse out.

Antibody Structure Questions

1. What is an antibody subclass?

Immunoglobulin classes (IgA, IgD, IgE, IgG, IgM in mammals) are defined by the structure of their heavy chains. Minor differences on heavy chains are found between subclasses of each class. Jackson ImmunoResearch offers polyclonal antibodies that are specific for mouse subclasses IgG1, IgG2a, IgG2b, IgG2c, and IgG3. We also have antibodies that are specific for camelid subclasses IgG 2+3. We currently do not have subclass-specific antibodies against other species.
Download Immunoglobulin Structure and Function Poster.

2. What are the differences between Fab, Fc, and F(ab')2 fragments?

These products are the result of proteolytic digestion of IgG. Digestion with papain results in Fab and Fc fragments, and digestion with pepsin yields F(ab’)2 fragments.

3. What is an antibody isotype?

Isotypes are distinct forms of antibody heavy and light chains, and may refer to class or subclass. Light chain isotypes are kappa and lambda, and heavy chain isotypes are alpha (IgA), delta (IgD), epsilon (IgE), gamma (IgG), mu (IgM). The heavy chains may be further differentiated as subclasses, e.g. gamma 1 (IgG1).

4. What is an isotype control?

Isotypes are distinct forms of antibody heavy and light chains, and may refer to class or subclass. Light chain isotypes are kappa and lambda, and heavy chain isotypes are alpha (IgA), delta (IgD), epsilon (IgE), gamma (IgG), mu (IgM). The heavy chains may be further differentiated as subclasses e.g. gamma 1 (IgG1).
An isotype control is a negative control that verifies specific binding of either a primary or secondary antibody. The isotype control is the same class or subclass as the experimental antibody, but is not specific for the antigenic target. Isotype controls may be purified IgG from normal serum of the species host of the experimental antibody; or in the case of monoclonal antibodies may be monoclonals of the same subclass, directed against an irrelevant antigen. Isotype controls for unlabeled antibodies are unlabeled, and controls for labeled antibodies are labeled with the same reporter molecule (fluorophore, enzyme, or biotin) as the experimental antibody.


Serological testing for diagnostics and disease surveillance

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Lateral flow tests for diagnostics

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Cross-adsorbed secondary antibodies and cross-reactivity

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Blocking WIth Fabs

Blocking: Use of unconjugated Fab fragments to block endogenous immunoglobulins and avoid off target signal   Background staining may be Read More...

Anti-Chicken Antibodies

Conjugated Anti‑Chicken antibodies are the detection partner for chicken‑derived antibodies, a host species gaining popularity as a component of Lateral Read More...

Anti-Human Secondary Antibodies

Jackson ImmunoResearch AffiniPure™ Anti-Human secondary antibodies are affinity purified polyclonal antibodies with specificity for human immunoglobulins. We offer Anti-Human secondary Read More...

Alexa Fluor Secondary Antibodies

Alexa Fluor® fluorescent dyes are widely recognized as superior fluorescent dyes available for conjugation. They are highly water soluble and Read More...

Silver Enhancement Protocol

Caution: Purity of water and cleanliness of glassware are crucial for optimal silver-staining results. Also, contact between metallic objects (for Read More...

Blocking and labeling with Fab fragments

Monovalent Fab Fragment Affinity-Purified Antibodies for Blocking and Double Labeling Primary Antibodies from the Same Host Species Monovalent Fab fragments Read More...

Applications

Common Applications of Secondary Antibodies Western Blotting Troubleshooting GuideWestern Blotting Western blotting with JIR secondary antibodies Western blotting is an Read More...

Secondary Antibody Conjugate Selection

Reporter Molecules Conjugated to Affinity-Purified Antibodies and Other Proteins For more information about the reporter molecules available conjugated to JIR Read More...

Imaging with VHH Antibodies

Using Whole Ig or Fab Fragment Secondary Antibodies Introduction Imaging with VHH antibodies (Single-domain antibodies, nanobodies†) is in its infancy, Read More...

Rapid Test Kits Development

Request a QuoteTechnical SupportPlace an Order Below is a selection of reagents for the development of COVID-19 rapid test kits. Read More...

JIR covid-19

Research Solutions for Coronavirus – Jackson Immunoresearch

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10 tips for better blots

Western blotting

Western blotting is a technique used to confirm the presence of target proteins and peptides from complex mixtures. Proteins are Read More...

How to select a Secondary Antibody

Choosing the right affinity-purified secondary antibody for your application The following information details how to use the Product Filter on Read More...

Mouse Subclass Specific Secondary Antibodies

Anti-Mouse IgG Subclass Specific Secondary Antibodies Mouse IgG subclasses Mice express 4 of the 5 available IgG subclasses making up Read More…

Light Chain Specific Secondary Antibodies

Western blotting after Immunoprecipitation For researchers who perform Western blotting following immunoprecipitation, antibodies specific for light chains or Fc fragments Read More…

Primary Antibodies for Signal Enhancement

Additional Antibody Specificities Anti-Biotin, Anti-Fluorescein and Anti-Digoxin antibodies Monoclonal Mouse Anti-Biotin, Anti-Fluorescein and Anti-Digoxin are available in a wide range Read More…

Cross-Adsorbed Secondary Antibodies

Cross-adsorbed (min X) secondary antibodies and cross-reactivity Immunoglobulins from different species share similar structures. Secondary antibodies raised against one species Read More...