Agarose bound Agaricus Bisporus Lectin (ABL)

AL-1423

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SKU Unit Size Price
AL-1423-2 2 ml
$262.00

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Description

Features:

  • Matrix is heat stable, cross-linked 4% agarose beads with a molecular exlusion of about 2x107 daltons
  • Bead diameter ranges in size from 45-165 microns
  • Matrix is stable in solutions at pH 3-11 as well as many organic solvents
  • Immobilized lectins are prepared using affinity purified lectins
  • Covalent attachment preserves lectin activity and minimizes conformational changes that might result in nonspecific or hydrophobic interactions
  • Hydrophilic spacer arm is inserted between the lectin and the matrix
  • Conjugated proteins are not leached off the beads by Tris or other routinely used buffers
  • No residual charges present after conjugation.  This minimizes non-specific binding to the matrix
  • Product supplied as a 1:1 suspension in buffer
  • 2 mg lectin/ml gel
  • Elution conditions: Fetal calf fetuin bound to Agarose Agaricus bisporus Lectin can be eluted with 0.1 M ammonium hydroxide. However, because glycoproteins vary in their structure, some glycoproteins may not elute under these conditions.

Specifications

More Information
Unit Size 2 ml
Applications Glycobiology, Affinity Chromatography
Matrix Conjugate Lectins
Sugar Specificity Gal(b-1,3)GalNAc (T disaccharide)
Conjugate Agarose

Documents

Product FAQs

I purchased an agarose bound lectin from Vector Labs. Do you have a protocol outline on how this may be applied in a column format?

Our agarose lectin products are supplied as hydrated matrix solutions in amber glass bottles. The agarose (bead) material will settle and you will see two phases in the tube supplied. The upper phase is buffer. A column can be prepared in a commercial plastic device such as Bio-Rad Cat # 732-6008 or an inverted Pasteur pipet with glass wool lightly packed in the neck to retain the agarose. 1) Draw (pipet) the desired amount of settled agarose-lectin (gel) from the stock bottle into the prepared column and let the buffer drain by gravity.(Sometimes an air bubble in the column tip prevents flow; tapping the column should get the flow started). 2) Wash the gel with 10 column volumes of buffer, such as HBS (10 mM HEPES, 0.15 M NaCl, pH 7.5) and discard the flow through. 3) Place a collection vessel (e.g. glass test tube) under the column tip and apply the glycoprotein-containing solution.Allow the solution to drain through using gravity. We recommend against pushing or pulling the material through the column. Retain the flow through material until the desired binding has been confirmed. 4) After sample application, wash column with 2-3 column volumes of buffer (or until the absorbance at 280nm is reduced to a satisfactory level) to remove unbound materials before elution. 5) Place a fresh collection vessel under the column tip.Apply the eluting solution again letting gravity do the work of moving the solution over the column. Note that in some cases, several column volumes of eluting solution may be required to achieved adequate release of bound material. 6) Following elution, the column can be prepared for reuse by washing with 10 column volumes of buffer. 7) If the column is to be stored, equilibrate the column with buffer containing 0.08% sodium azide. Cover the column with a plastic wrap, or similar, to prevent desiccation and keep at 4 degrees Celsius. The column will be stable for many months when stored under these conditions.

Technical Information

Agaricus bisporus lectin (ABL) isolated from the edible white button mushroom, is composed of two to four very similar isolectins, each a tetramer of approximately 17 kD subunits.  This lectin binds the Thomsen-Friedenreich antigen, also called T disaccharide, galactosyl (β-1,3) N-acetylgalactosamine.  Unlike peanut agglutinin, which does not bind sialylated T antigen, ABL binds either sialylated or asialylated forms.  Also, whereas peanut agglutinin has a proliferative effect on cancer cells, ABL is anti-proliferative, apparently without cytotoxicity.  ABL binds all blood groups and O-linked glycans of IgA subclasses.  Simple mono- and disaccharides are only marginally inhibitory.

Agarose bound* Agaricus bisporus lectin is prepared using our affinity-purified lectins. Heat stable, cross-linked 4% agarose beads with a molecular weight exclusion limit of about 2x107 daltons are used as the solid-phase matrix to which the lectins are covalently coupled. The attachment of the lectins to the beads is carefully controlled to preserve lectin activity and minimize conformational changes of the bound lectins that might result in nonspecific ionic or hydrophobic interactions. The technique we have developed to couple lectins to agarose beads inserts a hydrophilic spacer arm between the lectin and the matrix.

This coupling method provides several advantages over the traditional cyanogen bromide procedure:

  • Maximum carbohydrate binding activity of the coupled lectins is retained
  • Linkage is stable over a range of pH values
  • Conjugated proteins are not leached off the beads by Tris or other routinely used buffers
  • No residual charges are present after conjugation. This minimizes non-specific binding to the matrix.

Our agarose bound lectins are supplied at a constant concentration of lectin per ml of settled beads. The concentration for each lectin is selected to achieve the highest glycoconjugate binding capacity per mg of lectin present in the beads. Each lot is tested for its binding capacity using glycoproteins known to bind the lectin. This provides a guideline for the user and assures the quality of our agarose bound lectins.

Elution conditions:  Fetal calf fetuin bound to Agarose Agaricus bisporus Lectin can be eluted with 0.1 M ammonium hydroxide. However, because glycoproteins vary in their structure, some glycoproteins may not elute under these conditions.

*2 mg lectin/ml gel