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Journal of Clinical Microbiology, March 2000, p. 1267-1268, Vol. 38, No. 3
Department of Pathology and Laboratory
Medicine, University of California, Los Angeles, Santa Monica,
California 90402
Received 10 September 1999/Returned for modification 17 October
1999/Accepted 21 December 1999
The ColorPAC Giardia/Cryptosporidium (Becton Dickinson)
is a solid-phase qualitative immunochromatographic assay that detects and distinguishes between Giardia lamblia and
Cryptosporidium parvum in human stool. Agreement between
the Alexon-Trend ProSpecT Giardia Rapid EIA and the
ColorPAC assay was 166 of 172 (96.5%). Agreement between the
Alexon-Trend ProSpecT Cryptosporidium Rapid EIA and the
ColorPAC assay was 169 of 171 (98.8%). No cross-reactions were seen
with other parasites or human cells.
With the increasing interest in
potential waterborne outbreak situations, fewer well-trained
microscopists, and confirmation that both Giardia lamblia
and Cryptosporidium parvum can cause severe symptoms in
humans, laboratories are reviewing their options with regard to
immunoassay kits that can be incorporated into their routine testing
protocols (4, 14, 15, 20). Not only must these methods be
acceptable in terms of sensitivity and specificity but they must
provide clinically relevant, cost-effective, rapid results,
particularly in a potential waterborne outbreak situation (1, 2,
7, 13). Antigen detection assays for G. lamblia and
C. parvum have proven to be very useful in the diagnosis of
these infections (5, 10-12, 16-19). Advantages of these
assays include labor, time, and batching efficiencies that may lead to
reduced costs. These reagents offer alternative methods to the routine
"ova and parasite examination" (O&P) method and provide the added
sensitivity required to confirm infections in patients with low
parasite numbers.
Most commercially available immunoassays use the enzyme immunoassay
(EIA) format that requires multiple reagent additions, washing steps,
and incubations. A nonenzymatic rapid immunoassay for
Giardia and Cryptosporidium antigens has been
developed. This test (Genzyme Diagnostics Contrast
Giardia/Cryptosporidium) is marketed commercially
(Becton Dickinson ColorPAC Giardia/Cryptosporidium). The
assay can be performed in approximately 12 min on formalin-fixed (5 or
10% formalin or sodium acetate-acetic acid-formalin [SAF]) or
unfixed stool specimens. In the current environment of managed care and
cost containment, this new device may provide diagnostic testing that
is more accurate, rapid, and cost-effective than traditional methods.
Human fecal specimens were collected in 10% formalin or SAF and
polyvinyl alcohol-mercuric chloride base fixatives by the patients and
submitted to the laboratory from various clinics affiliated with the
UCLA Medical Center. Different parasites (nine protozoa, including both
trophozoites and cysts and 1 helminth; 74 positive specimens) and
stools containing human cells (21 positive specimens) were included in
the negative specimens; all specimens were patient specimens, not
seeded specimens. Specific organisms included Blastocystis
hominis (23), Chilomastix mesnili
(3), Dientamoeba fragilis (6),
Endolimax nana (14), Entamoeba coli
(6), Entamoeba hartmanni (3),
Entamoeba histolytica/E. dispar (10),
Iodamoeba bütschlii (5), Trichomonas
hominis (1), and Hymenolepis nana eggs
(3). Specific human cells seen in fecal specimens included
red blood cells (1), polymorphonuclear leukocytes
(19), and macrophages (3); these specimens also contained normal fecal debris, including various yeast cells. All
specimens were tested as indicated below; additional testing was
performed on discrepant specimens.
The following immunoassay diagnostic kits were used according to the
manufacturer's directions: (i) Alexon-Trend ProSpect Giardia Rapid EIA (Alexon-Trend-Seradyn, Ramsey, Minn.),
(ii) Alexon-Trend ProSpect Cryptosporidium Rapid EIA, (iii)
Alexon-Trend ProSpecT microwell EIA (for Giardia or
Cryptosporidium), and (iv) ColorPAC
Giardia/Cryptosporidium (Becton Dickinson, Sparks, Md.). The
Alexon-Trend-Seradyn rapid EIA methods were selected as being the most
comparable to the ColorPAC method in terms of time required for
performing the procedure. All immunoassay kits were used with unconcentrated, preserved stool specimens (5 or 10% formalin or SAF).
Specimens were tested using the following combinations: (i)
Alexon-Trend ProSpect Giardia Rapid EIA and ColorPAC
Giardia/Cryptosporidium or (ii) Alexon-Trend ProSpect
Cryptosporidium Rapid EIA and ColorPAC Giardia/Cryptosporidium. Discrepant results were tested
using the Alexon-Trend ProSpecT microwell EIA tests (for
Giardia or Cryptosporidium); this test was
selected in order to maintain consistency with a single manufacturer.
The ColorPAC Giardia/Cryptosporidium assay procedure
involves the addition of 2 drops of sample treatment buffer to a tube, the pipetting of 60 µl of uncentrifuged stool specimen into the tube,
the addition of 2 drops of a Giardia capture antibody
conjugate, and the addition of 2 drops of a colloidal carbon-conjugated
detection reagent for Giardia and
Cryptosporidium. After the sample is mixed, it is
immediately poured into the test device. Assay results are read after
10 min. Positive results are visualized as gray-black lines in the
appropriate position in the results window (Fig. 1). The tubes, pipettes, devices, and all
reagents are provided with the kit.
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Detection of Giardia lamblia and
Cryptosporidium parvum Antigens in Human Fecal Specimens
Using the ColorPAC Combination Rapid Solid-Phase Qualitative
Immunochromatographic Assay
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FIG. 1.
ColorPac devices demonstrating positive results: left,
positive control; middle, positive Giardia; right, positive
Cryptosporidium.
By the Merifluor Cryptosporidium/Giardia direct
fluorescent-antibody assay (the reference method), the O&P examination
(concentration, trichrome stain), and modified acid-fast staining
(concentration, modified acid-fast stain), 47 specimens were positive
for Giardia, 41 specimens were positive for
Cryptosporidium, and 256 were negative for both organisms.
Positive Giardia specimens (n = 47) and
negative specimens (n = 125) were tested using two
different kits. Agreement between the Alexon-Trend ProSpecT
Giardia Rapid EIA and the Becton Dickinson ColorPAC was 166 of 172 (96.5%); the six positives by microscopy, the Merifluor direct
fluorescent-antibody assay, and the ColorPAC were false negatives in
the Alexon-Trend ProSpecT Giardia Rapid EIA (Table
1). There were no cross-reactions with other organisms present in the fecal specimens; specificity was 100%.
Positive Cryptosporidium specimens (n = 41)
and negative specimens (n = 131) were tested using two
different kits. Agreement between the Alexon-Trend ProSpecT
Cryptosporidium Rapid EIA and the Becton Dickinson ColorPAC
was 169 of 171 (98.8%); the three negatives were false negatives using
the Alexon-Trend ProSpecT Cryptosporidium Rapid EIA. The one
false-negative C. parvum (ColorPAC) was confirmed by
immunofluorescence (Table 1). There were no cross-reactions with other
organisms present in the fecal specimens; specificity was 100%.
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In patients with giardiasis or cryptosporidiosis, the use of routine diagnostic methods such as concentration and trichrome or modified acid-fast staining may be insufficient to demonstrate the presence of these organisms (3, 4, 6, 9, 11, 21). Based on the O&P examination with trichrome stain, as well as the Meridian Giardia/Cryptosporidium Merifluor combination reagent, the sensitivity and specificity, respectively, using the ColorPAC device were as follows: G. lamblia, 100% and 100%; C. parvum, 97.6% and 100%. The Alexon-Trend ProSpecT Rapid EIAs did not perform as well, but results were within previous published expected values regarding sensitivity and specificity. The ability to concurrently detect and distinguish between Giardia and Cryptosporidium antigens in formalin-fixed or unfixed fecal specimens with a 10-min nonenzymatic immunoassay provides the user with another very useful diagnostic kit, the Becton Dickinson ColorPAC. The rapid immunoassays do not take the place of routine O&P examinations, but they are very useful when trying to confirm Giardia and Cryptosporidium infections (8).
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FOOTNOTES |
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* Corresponding author. Mailing address: UCLA Department of Pathology and Laboratory Medicine, 512 12th St., Santa Monica, CA 90402. Phone: (310) 393-5059. Fax: (310) 899-9722. E-mail: lgarcia1{at}gateway.net.
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REFERENCES |
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Journal of Clinical Microbiology, March 2000, p. 1267-1268, Vol. 38, No. 3
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
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