A. Introduction Intended use

The OSOM BV BLUE Test is an enzyme activity test for use in the detection of vaginal fluid specimens for sialidase activity, an enzyme produced by bacterial pathogens such as Gardnerella vaginalis, Bacteroides spp., Prevotella spp., and Mobiluncus spp. The OSOM BVBLUE Test is indicated for use in women suspected of having Bacterial Vaginosis (BV), e.g., women with vaginal discharge typical of BV and/or women with previous history of BV, as an aid in the diagnosis of BV infection. Test results should be considered in conjunction with other clinical and patient information (see Limitations of the Procedure).

Vaginos is one of the most common reasons that women visit obstetricians or gynecologists (1 – 3). The causative agents of this condition are bacteria such as Gardnerella vaginalis, Bacteroides spp., Prevotella spp. and Mobiluncus spp. Complications associated with BV include salpingitis, endometritis, post-hysterectomy infections, recurrent UTI’s, and an increased risk of PID and HIV(5 – 7). BV presents a serious danger in women, due to its significant association with placental infection, premature rupture of membranes, and preterm birth(8 –10). Studies have shown elevated sialidase activity in women with BV and an increased risk for preterm birth and low birth weight infants in patients exhibiting elevated sialidase activity (4, 11 – 15). The OSOM BV BLUE Test is designed to provide a clear, simple indication of elevated sialidase activity in patient vaginal fluid samples. The generation of a blue or green color indicates a positive test result; a yellow color indicates a negative test result.

The OSOM BV BLUE Test includes a chromogenic substrate of bacterial sialidase. In the test procedure, a vaginal fluid sample is placed in the BV Test Vessel. The sample then reacts with the chromogenic substrate. A Developer Solution is added after the reaction. If the sample has a high level of sialidase, a blue or green color will be seen in the BV Test Vessel or on the head of the swab. If the sample has no sialidase, or has very low levels, a yellow color will be seen in the BV Test Vessel.

1. Collect specimens with a swab from the lower one-third of the vaginal wall. Collecting specimens from the cervix should be avoided because:

a. it might increase risk to OB patients, and

b. cervical sialidase activity is usually higher than vaginal sialidase activity.

2. Do not use specimens from patients who have:

a. used a vaginal cream or ointment product,

b. douched, or

c. used spermicides, vaginal lubricants or feminine sprays within 72 hours of testing.

3. Test the patient specimen as soon as possible after collection.

4. If testing is not performed immediately, store the swabs either at room temperature for up to 48 hours or refrigerated for up to 7 days.

5. To transport patient specimens, place each swab in a clean, dry container such as a plastic or glass tube. Do not use any transport media. A false negative result may occur if insufficient sample is collected or the sample is obtained from a patient undergoing antimicrobial therapy.
C. Reagents and Supplies

. 1. Materials provided

a. 25 Test Vessels each containing 0.25 mg IBX-4041 component in 0.5 mL of an aqueous potassium acetate buffer solution (49mg/mL; 0.5 M; pH 5.5-6.0)

b. 1 Developer Solution Bottle containing 10.0 mL of an aqueous sodium hydroxide solution (40mg/mL; 1.0 M; pH>11.0)

c. Sterile Swabs

d. 1 Directional Insert

2. Materials required but not provided

a. OSOM BV BLUE Control Kit

b. Timer

3. Warnings and precautions

a. For in vitro diagnostic use only.

b. Do not use after the expiration date printed on the kit.

c. Do not store the kit at temperatures above 26°C (79°F).

d. Do not store the kit in strong light.

e. Follow laboratory safety guidelines in the collection, handling, storage and disposal of patient specimens and all items exposed to patient specimens.

f. Used tests should never be re-used.

g. This product is intended for vaginal fluid use only.

4. Storage and stability

a. Store the test kit at controlled temperature, 2° – 26°C (36°-79°F), out of direct sunlight. Store the test vessels inside the box. Kit contents are stable until the expiration date printed on the outer box.

NOTE: If temperatures in your facility may exceed 26°C (79°F), the kit should be refrigerated when not in use to ensure that the components remain stable until the expiration date printed on the packaging. Allow the kit to come to room temperature before running the test.
b. Store the control kit at 2° – 8°C (38°-46°F). Do not freeze. The kit is stable until the expiration date shown on the label.

5. Indications of instability:

a. Signs of possible product instability include:

1) A blue color in a BV Test Vessel when one drop of Developer Solution is added to the BV Test Vessel in the absence of a patient specimen.

2) Positive control does not give expected results.

3) Negative control does not give expected results.

Human specimens may harbor infectious agents. Use standard (universal) precautions when working with these materials including gloves and eye protection.

1. Internal Quality Controls. The OSOM BV BLUE Test contains two types of internal quality control with each test run. For daily quality control, the manufacturer recommends documenting these controls on each day of testing:

a. Type 1 Internal Control: Before adding a patient specimen, inspect the Test Vessel. It should contain a colorless liquid without precipitates (sediment). If the testing vessel contains a precipitate, the test is invalid. Do not use the Test Vessel.

b. Type 2 Internal Control: The OSOM BV BLUE Test has a two-color result format: blue/green is positive, yellow is negative. After running the test according to the instructions for use, the appearance of either a uniform yellow, blue, or green color in the testing vessel or a blue or green color on the swab assures proper mixing of the reagent and sample has occurred.

3. 
   Interpretation of results.
   
   1. 
      If the test fails to provide either a blue, green, or a yellow
      

2) Do not report patient results if either of the Internal Quality Controls does not produce expected results.

2. External Quality Controls. External Controls are used to test that the reagents are working properly. External Controls also verify that the test procedure is performed correctly.

a. A Control Kit that contains a positive control and a negative control may be purchased separately from Genzyme Diagnostics, Catalog No. 184.

b. Controls must yield expected results for both the positive and negative external controls before client samples may be tested and reported.

c. If QC testing fails:

1) Check expiration dates of the test kit and controls.

2) Ensure the instructions for testing were followed correctly.

3) Repeat the test.

d. If the controls still do not perform as expected, contact Genzyme Technical Service at 1-800-332-1042.

3. Frequency of external controls

a. Each week (Monday –Friday/Saturday) of testing prior to testing any client samples.

b. Each new lot

c. Each new shipment

d. Each new untrained operator.

F. Step by step instructions – External Controls

1. Remove the control kit from the refrigerator prior to use. Allow the control kit to warm to room temperature before use.

2. If the test kit is stored in a refrigerator, remove it prior to use. Allow the test kit to warm to room temperature before use.

3. Positive Control

a. Remove one test vessel and the developer solution from the test kit prior to use. Remove the cap from the test vessel.

b. Add one drop of BV Blue Positive Control the the BV test vessel.

c. Put the head of a swab from the test kit into the solution in the BV test vessel. Let the BV test vessel containing the swab stand for 10 minutes between 17˚ and 37˚C, (62.6˚ – 98.6˚F).

d. Add one drop of developer solution to the BV test vessel containing the swab. Gently swirl the mixture.

e. Read the results immediately after the developer solution.

4. Negative Control

a. Remove one test vessel and the developer solution from the test kit prior to use. Remove the cap from the test vessel.

b. Add one drop of BV Blue Positive Control the the BV test vessel.

c. Put the head of a swab from the test kit into the solution in the BV test vessel. Let the BV test vessel containing the swab stand for 10 minutes between 17˚ and 37˚C, (62.6˚ – 98.6˚F).

d. Add one drop of developer solution to the BV test vessel containing the swab. Gently swirl the mixture.

e. Read the results immediately after the developer solution.

5. Expected results

a. Positive Control Result: The positive control will produce a blue or green color in the test vessel and/or on the head of the swab.

NOTE: In the case of weak positive results, take the swab out of the vessel to determine in a blue or green color is present on the swab.

b. Negative Control Result: The negative control will produce a yellow color in the test vessel.

1. If the test kit is stored in a refrigerator, remove it prior to use. Allow the test kit to warm to room temperature before use.

2. Remove one BV Test Vessel and the Developer Solution Bottle from the kit prior to use. Remove the cap from the BV Test Vessel.

3. Collect a vaginal fluid sample with a swab. Contact the swab with the lower one-third of the vaginal wall. Collect as much fluid as possible.

4. Put the swab into the BV Test Vessel. Gently swirl the mixture.

5. Let the BV Test Vessel containing the swab stand for 10 minutes between 17˚ and 37˚C, (62.6˚ – 98.6˚F).

6. Add one drop of Developer Solution to the BV Test Vessel containing the swab. Gently swirl the mixture and read the results immediately.

1. There are two possible results:

a) Positive result: A blue or green color in the BV Test Vessel or on the head of the swab.

b) Negative result: A yellow color in the BV Test Vessel.

2. If the test fails to provide a blue, green, or yellow color result, the test is invalid.

3. A Positive Result shows a high level of sialidase activity.

4. A Negative Result shows a normal level of sialidase activity.

5. The OSOM BVBLUE Test can show sialidase activity in vaginal fluid at levels of ≥7.8U.
I. Procedural Notes

1. Read all instructions carefully before use. If a laboratory modifies the following test instructions including Quality Control, the test will be considered High Complexity and no longer considered Waived.

2. For in vitro diagnostic use only.

3. Do not use after the expiration date printed on the kit.

4. Do not store the kit at temperatures above 26°C (79°F).

5. Do not store the kit in strong light.

6. Follow your laboratory safety guidelines in the collection, handling, storage and disposal of patient specimens and all items exposed to patient specimens.

7. Used tests should never be re-used.

8. This product is intended for vaginal fluid use only.

9. Do not use samples from patients who have used vaginal cream products within 72 hours before testing.

10. Do not touch or collect fluid near the cervix.

11. In all clinical studies, no evidence of interference was observed for menses, blood, semen, birth control methods including birth control pills, Depo-Provera, Norplant, IUDs, condoms, or tubal ligation; or microorganisms including Staphylococcus, Streptococcus, E. coli, Candida albicans, Lactobacillus, among others.

1. Do not use samples from the cervix.

2. Patients may have mixed infections. The OSOM BV BLUE Test shows that sialidase enzyme is active in the sample. The OSOM BVBLUE Test does not show if other organisms such as yeast and parasitic organisms are present in the sample.

3. Test results should be considered in conjunction with other clinical and patient information.

4. Test operators must follow all instructions to:

a) collect the sample,

b) store the sample, and

c) use the test procedure properly.

If the instructions are not followed, the OSOM BVBLUE Test may not give correct results.
K. References:

1. Hillier, S.L. Diagnostic microbiology of bacterial vaginosis. Am. J. Obstet. Gynecol. 1993; 169:455.

2. Eschenbach, D.A. History and review of bacterial vaginosis. Am. J. Obstet. Gynecol. 1993; 169:441.

3. Hill, G.B. The microbiology of bacterial vaginosis. Am. J. Obstet. Gynecol. 1993; 169:450.

4. Briselden, A.N., et al. Sialidases (neuraminidases) in bacterial vaginosis and bacterial vaginosis-associated microfluora. J. Clin. Microbiol. 1992; 30:663.

5. Faro, S., et al. vaginal flora and pelvic inflammatory disease. Am. J. Obstet. Gynecol. 1993; 169:47.

6. Taha, T.E., et al. HIV infection and disturbances of vaginal flora during pregnancy. J. Acquired Imm. Def. Syndr. 1999; 20:52.

7. Royce, R.A., et al. Bacterial vaginosis associated with HIV infection in pregnant women from North Carolina. J. Acquired Imm. Def. Syndr. 1999; 20:382.

8. Chiam, W., et al. The relationship between bacterial vaginosis and preterm birth. A review. Arch. Gynecol. Obstet. 1997; 259:51

9. McGregor, J.A., et al. Premature rupture of membranes and bacterial vaginosis. Am. J. Obstet. Gynecol. 1993; 169:463.

10. Leitch, H., et al. Bacterial vaginosis as a risk factor for preterm delivery: A meta-analysis. Am. J. Obstet. Gynecol. 2003; 189:139.

11. McGregor, J.A., et al. Bacterial vaginosis is associated with prematurity and vaginal fluid mucinase and sialidase: Results of a controlled trial of topical clindamycin cream. Am. J.Obstet. Gynecol. 1994; 170:1048.

12. Cauci, S.,. et al. Immunoglobin A response against Garderella vaginalis hemolysin and sialidase activity in bacterial vaginosis. Am. J. Obstet. Gynecol. 1998; 178:511.

13. Andrews, A.A., et al. The preterm prediction study: failure of midtrimester cervical sialdase level elevation to predict subsequent spontaneous preterm birth. Am. J. Obstet. Gynecol. 1999; 180:1151.

14. Cauci, S., et al. Determination of Immunoglobin A against Gardnerella vaginalis hemolysin, and with prolidase and sialidase levels in women with bacterial vaginosis. J. Infect. Dis. 2002; 185:1614.

15. Cauci, S., et al. Determination of Immunoglobin A against Gardnerella vaginalis hemolysin, sialidase, and prolidase activities in vaginal fluid: Implications for adverse pregnancy outcomes. J. Clin. Microbiol. 2003; 41:435.

16. Nugent, R.P., et al. Reliability of diagnosing bacterial vaginosis is improved by a standardized method of Gram stain interpretation. J. Clin. Microbiol. 1991; 29:297.

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RL.57.01