Isolation of Sperm Cells
The isolation of single sperm cells is important for genetic fingerprinting to clarify questions regarding family relationships, for forensic medicine to uncover sexual offences as well as for in vitro fertilization processes.
Here we describe the isolation of single sperm cells using the CellCelector platform.
The unique combination of highly precise robot technology with inverted microscopy allows automating the detection as well as the isolation of single sperm cells using glass capillaries. Time consuming manual work may be replaced by standardized conditions using the CellCelector system.
Single sperm cells were observed and isolated in bright field illumination using a 20x-Objective.
The size of the sperm cells were determined on about 5 x 33 µm using some special software features for measuring objects microscopically.
For a better view the rectangular marked area is shown enlarged (bottom right).
The detection of single sperm cells may be done either in manual mode by mouse click on the live image screen or automatically using a set of adjusted detection parameters (diameter, area, elongation etc.) after sample scanning.
The basic detection is done by defining a range of gray values. Target cells can be separated from neighbor cells and from the background. Depending on how gray value thresholds were set, either entire single sperms or just the heads of the sperm cells were detected.
Set-up of parameters to detect automatically single human sperm cells.
Different ranges of gray values (A & B) allow the detection of different parts of the sperms (D and E, detected areas marked in green) based on the reference image (C). Considering additional parameters such as diameter, area, elongation (F), can improve the scanning result even more (G and H).
Borosilicate glass capillaries with an inner diameter of 30 µm were used to isolate single sperm cells. By developing an aspiration pressure right above the cell using a volume of 60 nl liquid, the cell was detached from the bottom and soaked with the liquid into the glass capillary. The isolated sperm cells were transferred into separate wells of 2x9-well Ibidi-slides. Each single cell isolation is automatically documented by images taken before and after the picking process, which are saved in databases.
Microscopic images were taken right before (left) and after (right) the isolation of a single sperm cell.
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