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Histology and Light Microscopy

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arrow13IHC Retriever Buffers

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Epitope Recovery Buffers

Epitopes of different nature may require different recovery buffers for the best results.

Most epitopes can be unmasked on FF/PE sections with R-Universal buffer (considering that the majority of commercial antibodies are prepared against peptide epitope or bacteria-expressed fragment of a protein). Nevertheless, we found some cases where buffers besides R-Universal achieve a better result. Depending on the structure of the epitope and amino acids comprising it, different buffers should be used for epitope recovery.

The type of epitope defines the most appropriate recovery buffer.

In cases where R-Universal may not be the best buffer, as well as standard EDTA etc., the Technical Data Sheets may be used to choose the most appropriate buffer for epitope recovery in relation to the type of the epitope.

In total, we offer 5 different buffers for antigen recovery. Most epitopes can be perfectly recovered in R-Universal buffer, including those usually recovered in Citrate/EDTA/Tris-EDTA buffers. Two new buffers offer solutions for rare, but still existing cases where antibodies give an incorrect pattern on a retrieved solution or no staining at all. We also now offer standard Low pH (Citrate) or "L-Recovery" and High pH "H-Recovery" (EDTA) buffers in case you already use antibodies known to work after tissue processing in one of these buffers.

62719-30 T-Universal Epitope Recovery Buffer (10x stock) 125 ml 77.00 Add to Cart
62719-35 T-Universal Epitope Recovery Buffer (10x stock) 500 ml 119.00 Add to Cart
62719-40 U-Universal Epitope Recovery Buffer (10x stock) 125 ml 110.60 Add to Cart
62719-45 U-Universal Epitope Recovery Buffer (10x stock) 500 ml 159.60 Add to Cart
62719-50 L-Universal Epitope Recovery Buffer (10x stock) 125 ml 75.60 Add to Cart
62719-55 L-Universal Epitope Recovery Buffer (10x stock) 500 ml 123.20 Add to Cart
62719-60 H-Universal Epitope Recovery Buffer (10x stock) 125 ml 75.60 Add to Cart
62719-65 H-Universal Epitope Recovery Buffer (10x stock) 500 ml 123.20 Add to Cart

Epitope Recovery Optimization Buffer Set

The 2100 Retriever allows the use of up to 6 different buffers in one cycle, each of the Slide Chambers accommodating one buffer. Each individual Slide Chamber can accommodate up to 18 slides. You can optimize the recovery buffer (and/or test the usability of buffer for recovery) for 18 individual antibodies in one go.

Such runs allow the establishing of the best/acceptable epitope recovery conditions for all antibodies that can later be used in experiments, which is especially important if you plan an IF multicolor staining of tissue sections.

We advise to do the primary test for individual antibodies with immunoenzyme staining, as this allows for better analysis of the pattern of staining and (eventual) non-specific staining of some areas/structures in section (i.e. blood vessels, etc.).

For your convenience, we offer a complete set of different recovery buffers for the tests, containing 125 ml of each 10x buffer stock.

62719-70 Epitope Recovery Set, includes 1 of each of the above Recovery Buffers 5x 125 ml 433.44 Add to Cart

Reduce The Background, Not The Specific Staining

Reliable time tested solution to unmask antigen on formalin-fixed sections.

Any antibody preparation has some potential to produce a non-specific reaction in the assay. This originates from:

  • Non-specific antibodies that are present in some proportion in any polyclonal antibody preparation, including affinity purified ones (often "affinity purified" means only isolation of IgG fraction on Prot A/G column, not the purification on the antigen column under very stringent conditions)
  • Low specificity antibodies among specific ones in polyclonal
  • Fragments of fallen apart IgGs in stored preparations, including monoclonal
  • Separate heavy and light chains of specific antibodies, produced by most hybridomas

All these are capable of binding non-specifically to molecules on tissue sections, blots, fixed cells and other objects for immune detection. In case of retrieved formalin sections the risk of non-specific reaction is even increased, since to active the epitope recovery the proteins comprising the tissue sections are denatured during HIER, thus making accessible many domains that are charged and are capable of binding the test immunoglobulins on non-specific manner.

The standard means to block non-specific binding of specific antibody preparation is to add some irrelevant protein, such as BSA, other serum, casein, etc. However, everyone who tried to do this knows that increasing (for effective blocking) concentration of such blocking agent leads to a great reduction of specific reaction as well. This is due to large blocking molecules binding to accessible sites on section and thus sterically blocking access of specific antibodies to epitopes of interest (schematically represented in the figure on the left, top). All our buffers developed for immune assays contain instead short (0.6-2 kD) peptides that are capable of block effectively non-specific reaction while not affecting the specific binding of antibody.

The presented collection of Immunohistology buffers has also some other benefits (see below) and allows you to achieve the best quality IHC result without compromising the antigen detection. The buffers can also be used in other immune assays, such as immunofluorescence on sections, flow cytometry on fixed cells, western blot, hybridization of sections with antibody detection.

The Retriever IHC buffers empower you to control non-specific staining on every step of immunohistochemistry. They are especially highly recommended for research pathology where, in contrast to diagnostics, many polyclonal and/or low-affinity antibodies are used.

All buffers available in 50 ml, 125 ml and 500 ml package. Ready to use.

sectionblockmsdsarrow11Section Block

A new class of blocking solutions based on chemically modified and fragmented ultra-pure casein. Effectively reduces unwanted binding of primary antibody and conjugates you use to charged surface of the slide and tissue section. Greatly reduces non-specific binding while preserving the specific reaction, by saturating potential non-specific protein-protein interactions. Moreover, in contrast to BSA-based, IgGm casein or serum - based blocking solutions there is no interaction of specific antibody and blocking protein itself. It is not comparable to other commercially available or home-made blocking solutions. Recommended for research and diagnostic pathology, especially for retrieved sections and polyclonal antibodies.

arrow11Antibody Diluent

Buffer for diluting your primary and secondary antibodies, especially if they were stored for a while, even at -20 in glycerol, or in refrigerator. Nonspecific binding of the antibodies, negative effects of disturbing substances and low or medium affinity cross-reactivities of the antibodies will be minimized, making your result more reliable. Excellent for IHC (frozen and formalin sections), flow cytometry on fixed cells, Western Blot and other immune assays.

When used in pathology, it also greatly reduces non-specific reactivity of human serum components and immunoglobulins in tissue, vessels and cells with mouse antibodies used on section.

For especially "trouble"-giving antibodies, as well as for in situ PCR applications, this dilutent may also be used as a washing buffer, preventing secondary binding of your analystes during washing.

arrow11HRP Conjugate Diluent

Specifically designed for preparing solution of your HRP-conjugate used as the detection reagent. It is the antibody-dilutent buffer with additional component for stabilizing your HRP-conjugate. Allows you to further standardize the assay preparing ready-to-use conjugate solutions in advance and store them in refrigerator without loss of activity.

Ordering Information

62710 Section Block   50 ml 159.00 Add to Cart
62711 Section Block   125 ml 278.00 Add to Cart
62712 Section Block   500 ml 1,200.00 Add to Cart
62713 Antibody Diluent msds 50 ml 172.00 Add to Cart
62714 Antibody Diluent 125 ml 375.00 Add to Cart
62715 Antibody Diluent 500 ml 1,400.00 Add to Cart
62713-01 Antibody Diluent for Frozen Sections 50 ml 172.00 Add to Cart
62714-01 Antibody Diluent for Frozen Sections 125 ml 375.00 Add to Cart
62715-01 Antibody Diluent for Frozen Sections 500 ml 1,400.00 Add to Cart
62716 HRP Conjugate Diluent msds 50 ml 172.00 Add to Cart
62717 HRP Conjugate Diluent 125 ml 392.00 Add to Cart
62718 HRP Conjugate Diluent 500 ml 1,410.00 Add to Cart
62718-15 Slide Washing Buffer msds 500 ml 135.00 Add to Cart

R-Universalmsdsarrow11R-UNIVERSAL Epitope Recovery Buffer (10x stock)

Why the Universal Buffer?

Epitope recovery on formalin-fixed, paraffin embedded tissue sections, requires heat-induced treatment in buffer or, sometimes, proteolitic treatment of the deparaffinized tissue section. Which buffer to use, greatly depends on the exact antibody and the properties of the recognized epitope, therefore, one can find in literature and practice use of many buffers, including Citrate pH 3.4, Citrate 6.0, EDTA 8.0, Tris 9.0- 10.0, Tris-EDTA, etc.

Moreover, for individual antigens also the time of recovery in the individual buffer should be defined, as this may be different, and the treatment often destroys the epitope.

EMS offers a novel (patent-pending) technology for epitope recovery, primarily based on reversing the fixation effects of formaldehyde, which created links primarily between ε-amino groups of lysine and other amino-groups.

The buffer was extensively tested in pathology departments in the United Kingdom, and has shown excellent results when used with different antibodies, including those that normally require for successful staining treatment only in Low, or only High pH buffers, or require protolithic treatment.

The EMS Universal Buffer may be used in any heat-induced epitope recovery system (the time and temperature of treatment should be tested), but was specifically adjusted for tissue sections processing in our Antigen Retriever (62700 Series).

Using our Universal Buffer in our Retriever guarantees the highest rate of success in recovering epitope for any antibody, especially one that was never previously used on formalin-fixed sections.

Properties

Clear, non-toxic solution.

Presentation

R-Universal Buffer is supplied as 10x concentrate. For epitope recovery dilute 1 part of stock with 9 parts of deionized water.

Application

For epitope recovery dilute 1 part of stock with 9 parts of deionized water.

Stability and Storage

The preparation is stable for 1 year when stored unopened at +4°C. Every lot is issued with a certificate indicating the expiry date.

After opening, store at +4°C in the refrigerator and use within 6 months.

Certification

Each lot is certified for compliance to specifications. The product is produced under DIN EN ISO 9001 :2008 Quality Management system for the products in Immunoassay Development and Measurement, Products for Bioanalytics and lmmunoassays.

Ordering Information

62719-10 R-UNIVERSAL Epitope Recovery Buffer (10x stock) 125 ml 139.00 Add to Cart
62719-20 R-UNIVERSAL Epitope Recovery Buffer (10x stock) 500 ml 229.00 Add to Cart

Antigen Retrievers for IHC arrow13arrow13