Cryptosporidium

Introduction

The fluorescent acid fast stain Auramine O (AuO) is simple and the widely used procedures for staining mycobacteria work well for Cryptosporidium. The present case study describes a new device for fluorescence microscopy, which can be employed on small microscopes at an affordable price. Furthermore it can be adapted on existing microscopes. Battery operation is also possible. Conventional light microscopy on Cryptosporidium smears usually requires the use of the 100× oil-immersion objective which causes prolonged observation times. Contrary to this, an Auramine O stained smear can easily be screened with a lower magnification objective in a fraction of the original time. The striking contrast between the brightly fluorescent oocysts and background facilitates the observation further.

FLUOLED® Easy: transmitted light fluorescence for the detection of Cryptosporidium

A unique, proprietary illumination system was developed using high-power solid-state (LED) light sources in transmitted light rather than reflected light. Sporozoites can be easily recognized inside the oocysts with the 40x, 50x, 60x dry or 100x oil-immersion objective, while artefacts stain homogenously and fluoresce less strongly.

case_cry_1 — FLUOLED® Easy on Olympus Cx41 microscope: the LED fluorescence module is designed to attach to a standard bright field microscope and is used in transmission mode.

case_cry_2

case_cry_3 — Sporozoites in Cryptosporidium oocysts stained with typical Auramine O staining patterns. Magnification: left: 20x, middle: 60x and right 100x. Excitation 450nm, emission long pass filter 500 nm.

Results

Transmitted-light fluorescence microscopy proved to be a valuable technology for observation of Auramine O stained Cryptosporidium. The use of a high power solid-state (LED) source enabled increased performance, significantly increased light source lifetime, reduction of initial costs and operating costs, and reduction of maintenance and heat production. Main FLUOLED® Easy product benefits are:

High contrast fluorescence images allow easier detection of parasites.
Light source lifetime: typically 30.000 hours, thus allowing many years of operation and cost savings.
It combines the advantages of high quantities of light transilluminating the sample with the high transmission values of the basic objectives found in this class of instruments.
No need of any special alignment procedure.
No warm-up time required for the light source.
Allows transmitted light bright field observation.
Battery pack option for field operation.

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