Speckle Counting
Speckle Counting - Related Videos
19:16
Live Cell Imaging of F-actin Dynamics via Fluorescent Speckle Microscopy (FSM)
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Cited by 8 •
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Selection, microinjection, and imaging of fluorescently-labeled F-actin via fluorescent speckle microscopy (FSM).
05:24
How to Build a Laser Speckle Contrast Imaging (LSCI) System to Monitor Blood Flow
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Cited by 25 •
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This video demonstrates how to build a Laser Speckle Contrast Imaging (LSCI) system that can easily be used to monitor blood flow.
10:19
Using a Hemacytometer to Count Cells
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2023
Many biomedical experiments require manipulation of a known quantity of cells, in order to achieve accurate, reproducible, and statistically-relevant data. Therefore, learning how to count cells is a particularly essential technique for any successful biomedical scientist. The most common way to count cells is by using a hemacytometer - an instrument that bears two laser-etched grids, which aid in the enumeration of an aliquot cells under a simple light microscope. This data can then be used to...
09:05
Transthoracic Speckle Tracking Echocardiography for the Quantitative Assessment of Left Ventricular Myocardial Deformation
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Cited by 9 •
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Speckle tracking echocardiography is an emerging diagnostic imaging technique for the quantitative assessment of global and regional myocardial performance. Standard view echocardiographic motion images are recorded and deformation parameters are subsequently measured by automated continuous frame-by-frame tracking and motion analysis of speckles within the B-mode images of the myocardium.
05:47
Counting and Determining the Viability of Cultured Cells
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Cited by 19 •
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Determining the number of cells in culture is important in standardization of culture conditions and in performing accurate quantitation experiments. In this video, we demonstrate how cells are counted using a hemacytometer.
FAQs
Background correction removes fluorescence that does not belong to discrete structures, while intensity thresholding sets the signal level used to separate candidate speckles from surrounding image background. Applying both steps helps prevent diffuse fluorescence from being mistaken for objects and makes subsequent segmentation or connected-component analysis more consistent across images.
Size and shape criteria act as filters after candidate objects are detected. They help exclude objects that are unlikely to represent the punctate structures being measured, reducing false counts caused by irregular or inappropriate image features. These criteria refine the object list before researchers summarize speckle number, distribution, or abundance.
Changes in counts can provide evidence that molecular structures have been reorganized or altered between cells or experimental conditions. Measurements of protein or RNA foci and nuclear bodies may reveal differences in molecular localization, organization, or abundance. The count is therefore useful as a quantitative indicator of cellular state.
A practical workflow begins with a biological microscopy image, corrects background fluorescence, and applies an intensity threshold to identify candidate speckles. Segmentation or connected-component analysis then separates individual objects. Finally, size and shape criteria help remove likely false detections, after which the remaining objects can be counted and compared.
The method can be applied to punctate biological signals such as protein foci, RNA foci, nuclear bodies, and other discrete fluorescent structures. Its value is greatest when the structures appear as separable spots in microscopy images, allowing their number, spatial distribution, or abundance to be quantified.
Comparisons should retain the distinction between how many speckles are present and how they are arranged or abundant. Counts can be compared across cells or experimental conditions, while distribution and abundance provide complementary measurements. Together, these readouts can show whether a change reflects altered molecular localization, organization, or broader cellular state.