Jackson ImmunoResearch supply secondary antibodies and proteins of the highest purity and specificity available. With a superb range of over 4000 products Jackson have whole IgG and (Fab`)2 affinity purified antibodies conjugated to AMCA, FITC, Cyanine dyes, TRITC, Rhodamine Red X, Texas Red, Biotin SP, Horseradish Peroxidase and Alkaline Phosphatase. For flow cytometry there are Phycoerythrin congugates and immunogold complexes for both light and electron microscopy. Having sold these reagents in the U.K. for over 18 years we can say they are simply the best!
Products Sub Categories
4nm LM Grade Colloidal Gold Complexes(Top)
We offer 4nm colloidal gold particles for light microscopy because this size may penetrate tissue better than larger particles.This size may also be used for electron microscopy in studies that require smaller particles since they are relatively uniform in size (coefficient variant less than or equal to 15%). However small aggregates are not removed from this grade by centrifugation in density gradients. Therefore, our 4nm particles should only be used for EM with the understanding that the possible presence od small aggregates may lead to misinterpretation of results. A detailed protocol for silver enhancement is provided with all orders for LM grade products. Includes Goat Anti Peroxidase colloidal Gold complex.
Affinity Purified Antibodies (WhoIe IgG) and Conjugates (Top)
All affinity purified antibodies are isolated from antisera by immunoaffinity chromatography using antigens coupled to agarose gels. A proprietory, sequential elution process is used to detatch purified antibodies from the solid phase antigen. All unconjugated affinity purified antibodies are supplied as sterile liquids without stabilizers or preservatives. All conjugated affinity purified antibodies are freeze dried with stabilizers and preservatives. Whole molecule IgG(H+L) reacts with both the Fc and F(ab`)2 portions of the IgG. Anti IgG(H+L) also reacts with other immunoglobulin classes (e.g. IgM, IgA,etc) since all immunoglobulins share the same light chains (either kappa or lambda).
Affinity Purified Anti Horseradish Peroxidase and Conjugates (Top)
Whole molecules of affinity purified anti horseradish peroxidase (HRP) may be used to detect HRP or to enhance signal by binding to any HRP conjugated molecule (such as HRP conjugated secondary antibodies or HRP conjugated streptavidin) or any HRP containing complex (such as PAP or HRP ABC).
Anti Fluorescein IgG Monoclonal Antibodies (Top)
IgG Fraction Monoclonal Mouse Anti Fluorescein and conjugates
Antisera to Enzymes, Serum Proteins, and Whole Serums (Top)
Polyclonal antisera from immunized hosts are lipid extracted to improve clarity, salt fractionated, dialyzed against phosphate buffer containing sodium azide and freeze dried. Antisera against whole serums are obtained by immunizing host animals with lipid extracted serums and chooseed serum fractions to obtain balanced, broadly reactive antisera. Antisera against whole IgG molecules (i.e. Anti IgG (H + L)) are recommended for bridging APAAP and PAP primary anti bodies.
Anti Subclass Specific Antibodies (Top)
Goat Anti Mouse Subclass Specific Antibodies in a variety of formats.
Biotin SP (long spacer) Conjugated Enzymes and Other Proteins (Top)
The Biotin SP (long spacer) conjugated proteins listed here are offered for the indirect bridged avidin biotin (IBRAB) method of Hsu et al. (J. Histochem. Cytochem. 1981. 29, 577). However, despite the possible advantages of this method, it may not be as sensitive as other biotin and streptavidin based methods (J.Histochem. Cytochem. 1989. 37, 1609; J. Histochem. Cytochem. 1992. 40, 135).
ChromPure Proteins (from Normal Sera) and Conjugates (Top)
ChromPure is our trade name for highly purified proteins from the serum of non immunized animals. The purified immunoglobulins in this section therefore do not represent antibodies directed against any known antigens. ChromPure proteins are prepared by a variety of methods, including ion exchange, gel filtration,dye ligand, metal affinity, and immunoaffinity chromatographies. F(ab`)2 (pepsin), Fab and Fc (papain) , and Fabµ and Fc5µ (trypsin) fragments are purified from enzyme digests of highly purified, whole molecules. No contaminating whole molecules or undesired fragments are observed at a minimum protein concentration of 20 mg/ml when tested by immunoelectrophoresis against anti whole serums, anti immmunoglobulins (class specific), or anti fragment specific antisera. However, these proteins are not guaranteed to produce monospecific antibodies when used for immunization. They are used primarily as experimental controls. All unconjugated ChromPure proteins are supplied without preservatives or stabilizers, either freeze dried or as sterile liquids in phosphate buffer. All conjugated ChromPure proteins are freeze dried in phosphate buffer containing preservatives and stabilizers.
EM Grade Colloidal Gold Complexes 6nm, 12nm or 18nm (Top)
EM grade products are distinguished from other commercial preparations by careful separation of monomeric particles from small agregates using ultracentrifugation in density gradients. The resulting monomeric colloidal gold protein complexes are suspended in sterile buffer containing preservatives and stabilizers. All particle sizes in the EM Grade category may be used for light microscopy and immunoblotting by those customers requiring larger particles.
F(ab`)2 Fragments of Affinity Purified Antibodies and Conjugates (Top)
F(ab`)2 fragments are used when binding of whole molecule, secondary antibodies to Fc receptors on cell surfaces needs to be avoided. As an alternative, binding of whole molecule, secondary antibodies to Fc receptors may be blocked by incubating cells at 4 degrees Centigrade in a buffer containing sodium azide and normal serum from the host species of the labelled secondary antibody. Please note that if a primary antibody is not a F(ab`)2 fragment it may also bind to Fc receptors and blocking with normal serum from the host species may not always be successful. CAUTION never block with normal serum or IgG from the host species of the primary antibody when using a labelled secondary antibody.
Fab Fragments for Blocking and Primary Antibodies from the Same Host (Top)
Monovalent Fab fragments of secondary antibodies can be used to sterically cover the surface of immunoglobulins for double labelling primary antibodies from the same host species, or to block endogenous immunoglobulins on cells or tissue sections.
IgG Free, Protease Free Bovine Serum Albumin (Top)
Bovine Serum Albumin (BSA) is used extensively as a carrier protein to dilute antibodies and a general protein blocking agent in immunoassays and immunodetection protocols. However many commercial preparations of BSA, even some of the highest purity grades, contain IgG that may become an antigen for cross reacting secondary antibodies. This is particularly common when using anti bovine IgG, anti goat IgG, anti horse IgG or anti sheep IgG, but may also show up with other antibodies that cross react with bovine IgG as well. The result may be loss of desired antibody activity, loss of antibody stability and/or increased background. Background may be derived from sticky soluble immune complexes or contaminating immune complexes or contaminating bovine IgG sticking non specifically and attracting cross reacting labelled secondary antibodies. Even a small percentage of contaminating IgG may create these problems due to the high concentrations of of BSA in many protocols. Jackson''s IgG free BSA, which is also protease free, should alleviate, many of these problems.
Light Chain Specific Antibodies (Top)
IgG Fraction of Monoclonal Mouse Anti Rabbit IgG, Light Chain Specific antibodies with a variety of conjugates.
Normal Sera and Gamma Globulins (Top)
Normal serums are lipid extracted, dialyzed, and freeze dried in phosphate buffer containing sodium azide. We strongly recommend the use of 4 5% normal serum, diluted in assay buffer, as a blocking agent to reduce background from non specific, conserved sequence, and/or Fc receptor bindings. Best results are obtained with diluted normal serum from the same host as the labeled antibody, as separate incubation steps before and after addition of the primary antibody. Gamma globulins are further purified from lipid extracted serum from non immunized hosts by salt fractionation and ion exchange chromatography. Gamma globulins are an inexpensive source of IgG with only trace amounts of other immunoglobulins and/or non immunoglobulin serum proteins. Gamma globulins are supplied as sterile liquids in phosphate buffer without any stabilizers or preservatives.
Peroxidase Anti Peroxidase (PAP) Soluble Immune complexes (Top)
PAP soluble immune complexes are prepared by the method of Sternberger et al. (J. Histochem. Cytochem. 1970. 18, 315). Theoretically, they consist predominantly of two anti horseradish peroxidase antibodies in soluble complex with three molecules of horseradish peroxidase. They are economical reagents for sensitive (Sternberger and Sternberger, J. Histochem. Cytochem. 1986. 34, 599) immunological detection of primary antibodies. Although higher sensitivity may be obtained with newer reagents such as horseradish peroxidase conjugated streptavidin (J. Histochem. Cytochem. 1988. 36, 317; and J. Histochem. Cytochem. 1989. 37, 1609), PAP continues to be used by some investigators, apparently due to its low non specific binding, which results in little or no background staining. F(ab`)2 PAP has been used to prevent the binding of PAP to Fc receptors. PAP soluble complexes are normally used at 25 50 g/ml. Whole antisera against IgG (H+L) are recommended for bridging PAP to primary antibodies. Antisera against IgG (H+L) or against the F(ab`)2 fragments of IgG will also bridge PAP to IgM primary antibodies (such as IgM monoclonal antibodies) by virtue of common light chain recognition. Normal serums from the same host species as the bridging antibodies are suggested as blocking agents to minimize non specific binding. All PAP soluble complexes are freeze dried at high concentrations for stability and high dilution capability. PAP soluble complexes, as well as other immunoperoxidase reagents, are not recommended for tissues or cells in which endogenous peroxidase activity is difficult to suppress. In such cases, other immunoenzyme reagents may be used. Alternatively, anti horseradish peroxidase conjugated with other enzymes or fluorophores may be used to enhance signal and reduce background since the final signal does not depend on the enzyme activity of peroxidase but on the antigenicity of horseradish peroxidase.
Phycoerythrin (PE) Conjugates (Top)
Phycoerythrins are among several kinds of light harvesting phycobiliproteins found in red, blue green and cryptomonad algae. They are highly fluorescent since each PE moleculae contains up to 40 fluorescent bile pigments and these pigments are arranged in such a way that there is minimal internal quenching. After conjugation to antibodies ther is little additional quenching which results in conjugates of high specific (fluorescence) activity compared with coventional fluorophore antibody conjugates.Phycoerythrins are attractive as immunfluorescent probes also because they can be excited by light over a wide range of the visible spectrum, are highly water soluble, have relatively low isoelectric points and lack potentially sticky carbohydrates.They have been used most successfully for flow cytometry and it should be noted that their high molecular weight obviates their widespread use in procedures reuiring good penetration in cells or tissues. We offer two forms of PE conjugates, B PE and R PE. Both forms can be excited by light at 488nm from an Argon laser and at 545nm from a mercury lamp. Since both forms of PE can be excited at the same wavelenght as fluorescein (488nm) but fluoresce at a longer wavelenght (580nm) than fluorescein (520nm) they have been used for double labelling in flow cytometry using a single argon laser.R PE, the form found in red macrophytic algae has been used more often for this purpose because it absorbs more light at 488nm than B PE, the form that we purify from the red microorganism, porphyridium cruentum. On the other hand, B PE absorbs more light at 545nm than R PE. B PE conjugates are available by request only.
Purified IgG Fraction of Monoclonal Mouse Anti Biotin and Conjugates (Top)
Our Monoclonal Mouse Anti Biotin offers another alternative for amplification procedures which can be used for in situ hybridization, immunohistochemistry and flow cytometry.
Purified IgG Fraction of Monoclonal Mouse Anti Digoxin and Conjugates(Top)
Monoclonal mouse Anti Digoxin may be used either as a direct conjugate or, for more sensitivity, unconjugated followed by a conjugated secondary antibody against mouse IgG. For fluorescence detection, Cy3 or Cy5 conjugates provide the highest sensitivity.
Solid Phase Immunoadsorbent Gels (Top)
ChromPure Proteins from the serum of non immunised animals coupled to cyanogen bromide activated 4% agarose gels for those wishing to prepare affinity purified antibodies or to remove cross reactive antibodies. Proteins are coupled at a concentration of Img/ml of settled gel.
Streptavidin and Conjugates (Top)
Streptavidin, a bacterial protein similar to egg white protein in its ability to bind biotin, has been used as a replacement for egg white avidin because of its more favorable chemical properties. It has almost no net charge at neutral pH, does not contain carbohydrate and exhibits lower non specific binding.Egg white avidin has a net positive charge at neutral pH and contains about 7% carbohydrate. Therefore, it may give higher background due to non specific binding to some cells or other surfaces.All conjugates of streptavidin and egg white avidin are recommended for use with Biotin SP (long spacer) conjugated, affinity purified, secondary antibodies and Biotin SP conjugated ChromPure proteins. Our research indicates that this system of covalently conjugated reagents is more sable, gives less background, and is more sensitive than the avidin biotin enzyme complex (ABC) method. This has been independently verified by Shi et al. (J. Histochem. Cytochem. 1988. 36, 317) and Milde et al. (J. Histochem. Cytochem. 1989. 37, 1609). The increased sensitivity may be due to enhanced tissue penetration and less steric hindrance since nominal molecular weights for all components of the conjugated streptavidin system are less than 200,000, which are considerably lower than those of ABC. Glucose oxidase conjugated egg white avidin has been used successfully in tissue sections and Western blots. It is not a mammalian enzyme, and thus it is recommended when background staining due to endogenous peroxidase or alkaline phosphatase is difficult to suppress in mammalian cells. The colored end product of glucose oxidase activity in the presence of tetrazolium salts is a non fading, insoluble formazan. This is particularly useful if fading or diffusion of peroxidase reaction products can not be prevented. In addition, glucose oxidase can be used with peroxidase or alkaline phosphatase for multiple labeling. We offer the following comprehensive list of fluorophores and enzymes conjugated to both streptavidin and egg white avidin for use in enzyme immunoassays, immunohistochemistry, flow cytometry, in situ hybridization, and immunoblotting procedures. Most streptavidin and egg white avidin products are freeze dried in buffer containing stabilizers and preservatives.
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