Why is FlowCam's Area-Based Diameter (ABD) More Accurate than Manual Microscopy When Measuring the Biovolume of Copepods?
Aquatic scientists have traditionally studied zooplankton to gain insights into aquatic systems' trophic status and ecological dynamics. Copepods, the most prevalent zooplankton group, connect primary producers with higher-level consumers.
Microscopy is widely regarded as the most precise and internationally accepted technique for measuring the size and biovolume of individual copepods. This method involves using a microscope to visually examine and measure the organisms, allowing scientists to gather detailed information about their physical characteristics. However, traditional microscopy often struggles to accurately account for protruding body parts, such as appendages and cilia. These structures can be difficult to measure precisely because they extend beyond the main body of the copepod and may not be fully visible or measurable in a single plane of focus.
As a result, biovolume estimations can frequently be inaccurate, as these important features are either underestimated or overlooked entirely. This limitation highlights the need for more advanced techniques that can provide a comprehensive and accurate assessment of copepod biovolume, considering all parts of the organism.
FlowCam has emerged as a preferred alternative to manual microscopy for researchers who aim to efficiently quantify plankton communities and assess their biovolume.
A Comparison of Biovolume Estimations
Pictured here: Biovolume estimations of various copepods using traditional microscopy (a), FlowCam ABD algorithm (b), and FlowCam ESD algorithm (c). Image from Karnan et al. (2017).
In the article "On the Accuracy of Assessing Copepod Size and Biovolume using FlowCam and Traditional Microscopy", published in the Journal of Geo-Marine Sciences, Karnan et al. (2017) compare FlowCam's ability to estimate length and biovolume to that of a traditional microscope. Microscopy has long been considered to be the most accurate and globally accepted method to measure the size and biovolume of individual copepods. However, protruding body parts (i.e. appendages, cilia, etc.) are not easily accounted for in biovolume estimations using traditional microscopy and often result in inaccurate estimations.
FlowCam is a flow imaging microscope that provides digital images of each particle, or in this case, copepod, and provides 40+ morphological measurements. For an Area Based Diameter (ABD) calculation, the diameter measured by the number of grey scale pixels of the binary image of copepods is converted to a circle with the same number of pixels. The total biovolume of the individual organism is generated from the pixel volume of the image.
ESD Method vs. Traditional Microscopy
For an Equivalent Spherical Diameter (ESD) algorithm, the mean of 36 diameter values measured at every 5-degree angle of the specimen is considered to estimate biovolume. This algorithm is accurate only when the specimens have a spherical or elliptical shape with short or no extended appendages. If the organism is not spherical and has extended appendages, it was found that FlowCam ESD-based biovolume overestimates due to the incorporation of appendages, and traditional microscopy underestimates due to the omission of appendages.
ABD Method vs. Traditional Microscopy
FlowCam's ABD algorithm generated the biovolume by selecting only the darkened region of the image. In all cases, it was found to be more accurate than ESD-based biovolume or biovolume calculated by traditional microscopy. The ABD algorithm of FlowCam provides a better estimation of copepod biovolume than traditional microscopy, as this method also incorporates the appendages and other extruded portions of the copepod. Due to the contribution of appendages and other extended body parts, ABD biovolume was found to be greater than biovolumes calculated using traditional microscopy.
Learn More
Read the entire paper here: On the accuracy of assessing copepod size and biovolume using FlowCam and traditional microscopy.
Reference:
Karnan, C., Jyothibabu, R., Manoj Kumar, T.M., Jagadeesan, L., Arunpandi, N. (2017) On the accuracy of assessing copepod size and biovolume using FlowCam and traditional microscopy. Journal of Geo Marine Sciences. Vol 46 (07), pp. 1261-1264.
