PARA-TEST PARASITE CONCENTRATION KIT

Technical Data #PT-50Va / 98.07.22

Diagnosis of intestinal parasites requires appropriate collection, processing and identification of protozoan cysts, helminth eggs and larvae in fecal material (1, 2, 3, 4, 5, 6, 7, 8). PARA-TEST is a complete standardized system for concentration and recovering protozoan cysts, helminth eggs and larvae from fecal material. PARA-TEST methodology is similar to the Ritchie formalin ether sedimentation concentration procedure (1, 2).

To achieve maximum recovery from preserved fecal material, the following components are utilized:

  1. Ethyl Acetate is used as solvent.
  2. 20% TRITON X-100 is added to preserved specimens containing mucus and/or fecal lumps. Acting as a surfactant, 20% TRITON-X-100 will free additionnal helminth eggs and parasites as adhesive forces are reduced in mucus and/or fecal lumps.
  3. /Filter device utilizes a consistent filter hole diameter to allow the passing of eggs, larvae and cysts while inhibiting the passage of macroscopic fecal matter and debris.
  4. Graduated polypropylene centrifuge tubes accomodate precise addition of preserved material and reagents, and are not soluble in ethyl acetate.
  5. Cotton tipped applicator sticks are used to clean the inside of the tube of remaining debris.


PRECAUTIONS
This medium is for IN VITRO diagnostic use only.


STORAGE
Store at room temperature (15-30 C).


LIMITATIONS
Feces for parasites examination must not be frozen and thawed. Formed specimens can remain satisfactory for a few hours at room temperature or overnight in a refrigerator. Soft and liquid specimens should be placed in fixatives within 1 hour. Specimen which cannot be fixed promptly should be either refrigerated or left at room temperature. Either saline or water can be used in the following procedure, but tap water will lyse Blastomyces hominis. Consult appropriate references for identification of fecal parasites. Observe package expiration date.

 

QUALITY CONTROL
Check reagent bottles for the expiration date. Do not use if expiration date has been exceeded. Examine bottle of 20% TRITON X-100 for cloudiness. Do not use if reagent appears cloudy. Visibly examine funnel/filter device for presence of plastic filter. Do not use if filter is absent or if filter spaces are blocked.

PROCEDURE
COLLECTION AND PREPARATION
Stool specimens preserved in 10% formalin (#FN-100) or Sodium Acetate-Formalin (#SAF-100) may be used. A portion of the specimen is added to the fixative in a ration of approximately 3 parts fixative to 1 part specimen and thorough mixed to break up any lumps. The stool/preservative mixture must stand for at least 30 minutes at room temperature to allow adequate fixation.
PROCESSING
Observe the stool/preservative mixture for mucus and/or fecal lumps.
If present, add 2 drops of 20% TRITON X-100 to the stool/preservative vial and shake vigorously.

  1. Place a funnel/filter device into a 15 ml centrifuge tube. Pour a sufficient volume of stool/preservative mixture through the funnel/filter such as that 1 ml of sediment will be deposited at the base of the centrifuge tube after initial centrifugation.
  2. Approximately 3 ml of stool/preservative mixture should be filtered for an heavy stool suspension, and up to 10 ml is required for a watery stool specimen. Do not force material through the tube. Centrifuge at 2500 rpm (650xg) for one minute.
  3. Decant the supernatant, retaining the fecal sediment. Suspend the fecal sediment in 8 ml of 10% formalin or SAF Fixative.
  4. Add 4 ml of ethyl acetate and place cap on the centrifuge tube. Invert centrifuge tube and shake vigourously for 30 seconds. Slowly remove cap to release pressure built up during shaking. Centrifuge the solution at 2000 rpm (500xg) for one minute. After centrifugation 4 layers should result as follows from the top down: Ethyl Acetate (approximately 3 ml), Plug of fecal debris and fat (approximately 0,5 ml), Discolored formalin or SAF (approximately 9 ml), Fecal sediment (approximately 0,5 ml)
  5. Holding the centrifuge tube vertically, free the plug of fecal debris and fat from the side of the tube by circling the rim of the layer with an applicator stick. Carefully decant the top 3 layers into a discard container. Continue to hold the centrifuge tube in the decanting position while thoroughly cleaning the inside of the tube of remaining debris with a cotton swab. This step is very important for lipid droplets which reach the sediment make examination difficult. Mix the remaining sediment with the small amount of fluid that drains back down from the sides of the tube.
  6. Within 30 minutes, transfer a portion of the fecal sediment to a clean slide and prepare the mount of choice for microscopic examination. If mounts are to be prepared later, a small amount of formalin may be added to the sediment and the tube stoppered.

 


REFERENCES

  1. Smith, J.W., and Bartlett, M.S. 1985 "Diagnostic Parasitology: Introduction and Methods" Manual of Clinical Microbiology. American Society for Microbiology. Washington, D.C. 4th ed. 595-608.
  2. Ritchie, L.S., 1948 An ether Sedimentation Technique for Routine Stool Examination. Bull. U.S. Army Med. Dept. 8:326.
  3. American Society for Parasitology 1977 Procedure suggested for use in examination of Clinical specimens for Parasitic Infection. J. Parasitol. 63:959-960.
  4. Burrows, R.B., 1965 "Microscopic Diagnosis of the Parasites of Man". Yale University Press, New Haven.
  5. Garcia, L.S., and Shimizu, R. 1981 Comparaison of Clinical Results for the use of Ethyl Acetate and Diethyl Ether in the Formalin-Ether-Sedimentation Technique performed on Polyvinyl Alcohol Preserved Specimens. J. Clin. Microbiol. 13:709-713.
  6. Garcia, L.S., and Vogue, M., 1980 Diagnostic Clinical Parasitology: 1. Proper Specimen Collection and Processing. Am. J. Med. Technol. 46:459-467.
  7. Melvin, D.M. and Brooke, M.M., 1980 Laboratory Procedures for the Diagnosis of Intestinal Parasites. U.S. D.H.E.W. 80:8202. CDC Atlanta, Ga.
  8. Young, K.H., Bullock, S., Melvin, C.M., and Sprull, C.L., 1979 Ethyl Acetate as a Substitute for Diethyl Ether in the Ether-Formalin Sedimentation Technique. J. Clin. Microbiol. 10:852-853.
  9. Markell, E.K. and Quinn, P.M., 1977 Comparaison of immediate Polyvinyl Alcohol (PVA) Fixation with delayed Schaudinn's fixation for the demonstration of protozoa in stool specimens. Am. J. Trop. Med. Hyg. 26:1139-1142.
  10. Smith, J.W., and Bartlett, M.S., 1985 Diagnostic Parasitology: Introduction and Methods. p.595-611 In Lennette, E.H. and al. Eds. Manual of Clinical Microbiology 4th ed. American Society for Microbiology Washington, D.C.


CATALOGUE NUMBERS

PARA-TEST KIT

PT-50

This kit contains:

50 Funnel/Filter devices
50 Polypropylene centrifuge tubes, 15 ml
50 Centrifuge tubes caps, plug type
50 Cotton tipped applicator sticks
15 ml 20% TRITON X-100 Reagent
225 ml Ethyl Acetate Reagent

 

 

PARA-TEST KIT

PT-50V

This kit contains:

50 Funnel/Filter devices
50 Polypropylene centrifuge printed tubes, 15 ml
50 Centrifuge tubes screw caps
50 Cotton tipped applicator sticks
15 ml 20% TRITON X-100 Reagent
225 ml Ethyl Acetate Reagent


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