Shreffler:Periodate Treatment

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Overview

Periodic acid can be used to degrade carbohydrate structures in order to test for the role of glycans in biological assays.

Materials

Procedure

Treatment of soluble protein:

  1. Bring CPE to ~3 mg/mL in 1 mL PBS
  2. Add 1 ml 100 mM sodium metaperiodate in PBS (pH 7.2)
  3. Incubate at 4°C in dark for 18h (O/N)
  4. Add 1 ml 260 mM glycerol
  5. Dialyze against PBS
  6. Protein assay and gel (approx protein conc 1 mg/mL

Treatment of plate bound protein (from reference 2):

  1. Coat wells with CPE per routine
  2. Add 0.25 mM sodium acetate buffer, pH 4.5 with 10 mM sodium metaperiodate
  3. Incubate 2h at 37°C in the dark
  4. Wash 1X with dH2O
  5. Add 0.5M sodium borate buffer, pH 8.0, containing 0.5 M Sodium borohydride for 30 min at 25°C
  6. Use immediately

Discussion

discuss this protocol

References

  1. Sato C, Inoue S, Matsuda T, and Kitajima K. Development of a highly sensitive chemical method for detecting alpha2-->8-linked oligo/polysialic acid residues in glycoproteins blotted on the membrane. Anal Biochem. 1998 Aug 1;261(2):191-7. DOI:10.1006/abio.1998.2718 | PubMed ID:9716421 | HubMed [Sato]
  2. Hino S, Matsubara T, Urisu A, Aoki N, Sato C, Okajima T, Nadano D, and Matsuda T. Periodate-resistant carbohydrate epitopes recognized by IgG and IgE antibodies from some of the immunized mice and patients with allergy. Biochem Biophys Res Commun. 2009 Mar 13;380(3):632-7. DOI:10.1016/j.bbrc.2009.01.138 | PubMed ID:19285013 | HubMed [Hino]
All Medline abstracts: PubMed | HubMed


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