Shreffler:Periodate Treatment

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

Materials

 * 15 mL polypropylene tubes
 * PBS
 * 50 mM Sodium Acetate Buffer, pH 4.5
 * 100 mM Sodium Borate Buffer, pH 8.0
 * Sodium Metaperiodate: |SIAL&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC Sigma cat no. S1878
 * Glycerol
 * Sodium Borohydride: |ALDRICH&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC Sigma cat no. 480886

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
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Contact

 * Who has experience with this protocol?
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