Sample preparation of milk (liquid).

Modified on Tue, 3 Oct, 2017 at 10:25 AM

This is a suggested sample preparation method for milk (liquid).

NOTE: The volume of supernatant used in the assay can be adjusted to suit the sensitivity of the assay being used.

SAMPLE PREPARATION (Liquid Milk):

1. To a 1.5 mL polypropylene microfuge tube, add the following with mixing after each addition:

Pipette into microtube	Volume
sodium phosphate buffer (0.4 M)	0.20 mL
Distilled water	0.70 mL
milk sample	0.50 mL
concentrated Carrez II solution	0.05 mL
concentrated Carrez I solution	0.05 mL

Mix by vortex and then centrifuge at 13,000 rpm for 10 min.

Carefully pipette the clear supernatant for use in the assay.

PREPARATION OF REAGENTS:

1. Sodium phosphate buffer (0.4 M, pH 7.6, 4 mM magnesium sulphate): 500 mL
Add 24 g of Na₂HPO₄ (Sigma cat. no. 71640), 4.3 g of NaH₂PO₄.
H₂O (e.g. Sigma cat. no. 71506) and 0.5 g of MgSO₄.7H₂O (e.g. Sigma cat. no.
M5921) to 450 mL of distilled water. Adjust the pH to 7.6 with
1 M NaOH (40 g/L) and make the volume up to 500 mL with distilled
water. Store at 4°C.

2. Concentrated Carrez I solution: 200 mL
Dissolve 30 g of potassium hexacyanoferrate (II) {K₄[Fe(CN)₆].3H₂O}
(e.g. Sigma cat. no. P9387) in 200 mL of distilled water. Store at room
temperature.

3. Concentrated Carrez II solution: 200 mL
Dissolve 60 g of zinc sulphate {ZnSO₄.7H₂O} (e.g. Sigma cat. no. Z4750)
in 200 mL of distilled water. Store at room temperature.