POTENTIAL OF FLOW CYTOMETRY FOR ROUTINE ALGAL COUNTING AND DETECTION OF CYANOBACTERIA
Report No FR0289

M M I Harman

March 1992

SUMMARY

I OBJECTIVES

The objectives of this work were:

1. To assess by flow cytometry the difficulties of analysing algal species which normally form long chains or clusters;

2. To investigate the possibility of finding optical properties which would serve to identify specifically Cyanobacteria;

3. To evaluate the optical and fluid flow design constraints in order to draft a specification for a practical low-cost portable flow cytometer for phytoplankton analysis.

In addition, the possibility of adapting an existing low-cost flow cytometer was investigated.

II REASONS

The production of toxins from blue-green algae can lead to problems with water used for potable supplies and recreational activities. Conventional techniques for the identification and counting of algae are both labour intensive and time consuming and during periods of increased sampling requirements can stretch the resources of microbiological laboratories and impose severe restrictions on existing monitoring programmes.

III CONCLUSIONS

The signal derived from some species which have a distinctive filamentous morphology, e.g. Anabaena, suggests that individual cells could be counted.

Emission spectra obtained for each of several species of green algae and Cyanobacteria show marked differences.

Although the measurement of Cyanobacteria is currently technically feasible, none of the compact flow cytometers currently available can be used without extensive modification. Constraints on design, imposed by the size and power requirements of current laser light sources, mean that at present the development of a portable low-cost flow cytometer for this application is not a practical proposition and any instrument developed in the immediate future would be restricted to the laboratory.

However, the requirement for a rapid optical method for routine counting of green algae could possibly be met by a portable arc-lamp illuminated flow cytometer because the high sensitivity to very small cells is not required.

IV RECOMMENDATIONS

Before continuing further with the development of a low-cost portable flow cytometer it is recommended that the potential users of the technique be consulted to establish their requirements for such an instrument. In view of the likely restriction of the technique for the detection of Cyanobacteria to the laboratory for the immediate future, the requirements for such an instrument should be assessed carefully before pursing this line of study further.

Further work is recommended on the development of techniques that could be applied to existing flow cytometer designs, thereby offering increased speed and sensitivity over existing methods, e.g. determination of bacterial species by immunofluorescent staining or the detection of pathogens in natural waters using fluorescence-labelled antibodies.

V RESUME OF CONTENTS

This report details the work carried out by Strathclyde University and discusses the possibilities of modifying a low-cost flow cytometer which has been designed for use in biomedicine.

The report presents the methods investigated and results obtained.

A sample preparation technique that may enable simple flow cytometers to be used for routine algal counting is described. A method for comparison of emission spectra from green algae and Cyanobacteria is described, which may form the basis of a technique for discrimination.

Design criteria for a flow cytometer for the analysis of algae are reported, including the flow cell design, type and configuration of the optics. The use or modification of existing flow cytometers to meet the requirements of phytoplankton analysis is discussed.

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