Alkaline hemolysis fragility is dependent on cell shape: Results from a morphology tracker

Abstract

Background: The morphometric analysis of red blood cells (RBCs) is an important area of study and has been performed previously for fixed samples. We present a novel method for the analysis of morphologic changes of live erythrocytes as a function of time. We use this method to extract information on alkaline hemolysis fragility. Many other toxins lyse cells by membrane poration, which has been studied by averaging over cell populations. However, no quantitative data are available for changes in the morphology of individual cells during membrane poration-driven hemolysis or for the relation between cell shape and fragility. Methods: Hydroxide, a porating agent, was generated in a microfluidic enclosure containing RBCs in suspension. Automatic cell recognition, tracking, and morphometric measurements were done by using a custom image analysis program. Cell area and circular shape factor (CSF) were measured over time for individual cells. Implementations were developed in MATLAB and on Kestrel, a parallel computer that affords higher speed that approaches real-time processing. Results: The average CSF went through a first period of fast increase, corresponding to the conversion of disco-

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“…Small ions and associated water molecules enter the cell, creating a positive osmotic pressure, resulting in swelling and subsequent bursting of the cell, a process referred to as colloid-osmotic lysis (Pooler, 1985). Hemolysis in the presence of detergent depends intrinsically on the osmotic fragility of erythrocytes (Ionescu-Zanetti et al, 2005), a property that depends on erythrocyte shape, membrane flexibility and cell surface area-tovolume ratio (Stuart and Nash, 1990). Flexibility of the cell membrane is also one of the major factors determining erythrocyte deformability (Srour et al, 2000).…”

Finding the Next Computational Model: Experience with the UCSC Kestrel

“…We have shown that with our next processor board generation with twice as many of the original 64-PE chips and a pipelined controller enabling a doubling of clock speed, Kestrel 2 (Fig. 14), the speedup would enable almost-real-time processing [22].…”

“…13) [22]. The application also enhances the video resolution to be able to accurately measure the cells_ parameters needed for the experiment-area, perimeter, and sphericity.…”

High-Throughput Assessment of Cellular Mechanical Properties

“…These early osmotic fragility tests characterized the quantity of cells lysed at a given time using bulk absorption measurements of hemoglobin or measures of intact pellet size after centrifugation. More recently, individual cells undergoing lysis have been tracked using video microscopy to study osmotic fragility in RBCs in a static setting (145). The approaches taken to increase the throughput of osmotic fragility tests make use of microfluidic devices that create osmotic shock by either electroporation (146) or exposure to a hypotonic solution (147).…”