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Ovucheck Plasma (Canada)

This page contains information on Ovucheck Plasma for veterinary use.
The information provided typically includes the following:
  • Ovucheck Plasma Indications
  • Warnings and cautions for Ovucheck Plasma
  • Direction and dosage information for Ovucheck Plasma

Ovucheck Plasma

This treatment applies to the following species:
Manufacturer: Biovet

Measurement of Progesterone in Plasma or Serum

Insert

2013-06-19

The OVUCHECK® PLASMA ELISA kit is an immunoenzymatic test which provides a simple, reliable and precise measurement of progesterone in plasma or serum from a number of species (for example cows and bitches.

Each kit contains sufficient reagents for up to 92 tests plus four standards.

OVUCHECK® PLASMA is used for oestrus detection and assessment of pregnancy status and luteal function.

Principle Of The Test

The OVUCHECK® PLASMA test is based on the competitive binding of unlabelled progesterone present in the standard or sample, and a fixed quantity of progesterone labelled with the enzyme alkaline phosphatase (AP) (conjugate), to binding sites on a limited amount of specific progesterone antibodies.

The wells are pre-coated with specific progesterone antibodies, providing a solid phase for the capture of the progesterone present in the samples, the standards or the conjugate. After incubation, all components other than those bound to the plate wells are washed away.

The amount of bound AP-labelled progesterone remaining on the wells is inversely proportional to the concentration of the unlabelled progesterone present in the sample. The bound labelled progesterone is then measured by making the AP react with its substrate during a second incubation.

The colour produced is measured spectrophotometrically and the concentration of progesterone in the sample is determined from a standard curve.

Precautions

- Store the kit at 2-7°C. DO NOT FREEZE.

- Do not use the kit after the expiry date indicated on the package.

- Do not mix the reagents from different lot numbers.

- The standards and the conjugate contain a preservative. When emptying the wells’ contents into a sink, thoroughly flush away with an excess volume of tap water.

- For in vitro veterinary diagnostic use only. The components or their residues must not be allowed to come into contact with livestock.

- Do not pipet by mouth.

- If eyes or skin are splashed, wash thoroughly with tap water.

- The material used in this kit must be considered as infectious. Therefore, all waste must be decontaminated before being discarded.

- Dispose of the substrate and the stop solution according to local regulations for chemicals.

Material

Reagents provided with the kit:

Components

Quantity

- 12 strips of 8 wells coated with progesterone antibodies

1

- Ready-to-use progesterone standards (Each of: 1.0; 2.5; 5.0; 10.0 ng/mL)

1.0 mL

- Ready-to-use conjugate-(Prog.-AP)

26 mL

- Substrate buffer

25 mL

- Substrate tablets

3 X 40 mg

- Ready-to-use stop solution

20 mL

- Graph paper

1

Materials Required but not Provided:

- Purified water

- Adjustable single- and multi-channel micropipettes

- Single-use micropipette tips

- ELISA 96-well microplate reader equipped with 405 nm filter

Execution

A. Substrate Preparation

Add 3 substrate tablets to the substrate buffer and shake it until complete dissolution.

If all the substrate is not required immediately, it can be dispensed into clean plastic containers and stored at 2-7°C for 1 week or aliquoted and frozen for 3 months at -20 ± 4°C. Keep the substrate away from light until ready to use.

B. Sample Collection

Plasma:

- Blood samples should be collected into heparinised collection tubes (do not use EDTA tubes).

- Centrifuge the blood sample within 30 minutes of collection and draw off the plasma into a clean container. The plasma sample can be kept in the refrigerator for up to one week before testing or in the freezer for many months.

- If centrifugation is not immediately available, keep the blood sample as cool as possible and centrifuge within 24 hours of collection. Progesterone levels fall by as much as 30% under these conditions.

Serum:

- Blood samples should be collected into a non-heparinised tube (do not use serum separator tubes). Once the clot has formed, centrifuge the tube within 30 minutes and draw off the serum into a clean container.

- If a centrifuge is not available, the serum should be drawn off as soon as the clot has settled. Serum samples can be kept in the refrigerator for up to one week before testing, or in the freezer for many months.

C. Test Procedures

1. Bring all components to room temperature before use (22 ± 3°C) before use (approximately 30 minutes).

2. Homogenize samples and reagents just before use.

3. Free the microtitration plate from the protective packaging. Take the quantity of microwells needed for the test (4 wells for the controls and 1 well per sample to be tested). Wells can be separated from one another. Return unused microwells strips to storage at 2-7°C.

4. To prevent any mistake, it is recommended to identify the wells used by making a schematic representation.

5. It is important to distribute samples and reagents in the wells in the minimum amount of time required (less than 10 minutes) to ensure a similar reaction time in all wells. If there are too many samples test them in smaller batches.

6. Add 10 µL of each four standards to be used to the appropriate wells.

7. Add 10 µL of each sample to be tested to the appropriate wells.

8. Immediately add 200 µL of conjugate to every used well. Take care not to contaminate the micropipette tips with the samples. Gently shake the wells to homogenize reagents.

9. Incubate for 30 minutes at room temperature away from light.

10. Empty wells and gently wash them with a pipette or a wash bottle using purified water at room temperature. Repeat twice more. Tap dry on absorbent paper. Do not let the wells dry completely.

11. Add 200 µL of substrate (see section A) to all wells.

12. Incubate for 30 minutes at room temperature away from light.

13. Add 100 µL stop solution to all wells.

14. Read the results at 405 nm. The reading should be done no later than 15 minutes after the addition of the stop solution. See « RESULTS » section.

15. At the end of testing, return components to storage at 2-7°C.

Interpretation

Results

Set spectrophotometer or plate reader to read absorbance at 405 nm. Zero instrument on air. Draw a standard curve by plotting standard absorbance values on the graph paper provided or by using appropriate software (e.g. Excel). The progesterone concentration of the samples can then be read from the curve. An Excel spreadsheet is available on Biovet website at: www.biovet.ca/kits.

Standard Curve Example

P4 standard (ng/ml)

OD (405 nm)

1

2.141

2.5

1.507

5

0.758

10

0.459

Guidelines for the detection of estrus

Species

Progesterone (ng/mL)

Suggested day of sampling for testing

Cow

< 2

19

Bitch: follow-up of blood progesterone concentration to predict ovulation day and optimal breeding day

Progesterone (ng/mL)

Estrus cycle phase

Interpretation

< 3

Proestrus

Retest in 2 days.

3-10

Estrus

Retest in 1 or 2 days.

> 10

Luteal phase

Mate within 48 hours after the progesterone level reaches 10 ng/mL.

Vaginal cytology:

- If predominance of cornified epithelial cells, mate immediately.

- If predominance of intermediate and parabasal cells, it is too late to mate.

Specificity

The interference (cross-reactivity) from steroids other than progesterone is insignificant (less than 1%) except for:

11α-Hydroxy-progesterone

66.0%

5-Pregnan-3β-ol-20-one

16.0%

Deoxy-corticosterone acetate

3.0%

5β-Pregnan-3, 20-dione

4.5%

5α-Pregnan-3, 20-dione

3.3%

Biovet Inc. 4375, Beaudry ave, St-Hyacinthe (Qc) J2S 8W2 Canada

Tél.:1-888-8BIOVET Fax: (450) 771-2996 Technical services: support@biovet-inc.com www.biovet.ca

NAC No.: 36940005

BIOVET INC.
4375 AVENUE BEAUDRY, ST-HYACINTHE, QC, J2S 8W2
Telephone:   450-771-7291
Toll-Free:   1-888-8BIOVET
Order Desk:   1-888-824-6838
Technical Service:   1-888-824-6838
Fax:   450-771-2996
Website:   www.biovet.ca
Email:   support@biovet-inc.com
Every effort has been made to ensure the accuracy of the Ovucheck Plasma information published above. However, it remains the responsibility of the readers to familiarize themselves with the product information contained on the Canadian product label or package insert.

Copyright © 2016 North American Compendiums. Updated: 2016-10-31

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