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New Point-of-Care Method for Leukocyte Differential Count

Jan 28, 2014 7:58:23 PM / by Maria

Independent study uses human venous whole blood from HemaCare to investigate the leukocyte differential count

Leukocyte differential count is one of the most frequently ordered clinical tests in hospitals. This test enumerates total leukocyte cells in blood and the differential count of four leukocyte subtypes (lymphocytes, neutrophils, monocytes, and eosinophils) for diagnosis of diseases such as: infections, inflammations, allergies, etc.

In recent years, there has been an emerging need for a point-of-care test of leukocyte count. Situations such as out-patient monitoring, medical care in remote areas, and telemedicine, are driving this need. However, the leukocyte count test is mainly performed in central clinical laboratories, which require bulky instruments and consume large amount of reagents that are not suitable for point-of-care applications.

By implementing a fluorescent dye assay on a sheathless microflow cytometer, researchers at California Institute of Technology  reported a promising approach to miniaturized cell analysis as an optimal method for point-of-care applications [1].

To solve this problem, there are increasing efforts to miniaturize the leukocyte count test assays. Yu-Chong, Tai and his team of researchers recently published their findings in the " Lab On A Chip" journal about " Four-part leukocyte differential count based on sheathless microflow cytometer and fluorescent dye assay". Using human whole blood samples from healthy donors, supplied by HemaCare, the researchers investigated the protein expression profile of leukocytes (lymphocytes, neutrophils, monocytes and eosinophils) in a microflow cytometer. They did so by applying the flow cytometry principles of labeling followed by detection:

(1) Labeling: target cells are labeled with fluorescent dyes (rather than  fluorescent-conjugated-antibodies) which provides increased stability  detecting target cells by corresponding optical signals (fluorescence).  Three fluorescent dyes: (PI), (FITC) and (BO21) introduce a high intensity of fluorescence to selectively distinguish leukocytes from other blood cells.

(2) Detection: leukocytes are counted as the sample passes through a sheathless, microfluidic channel.  

By confining the blood sample into a narrow stream by the microflow sheathless channel, the leukocytes were individually counted. This set-up reduced the chance of two or more leukocytes flowing through the detection channel together. The assay detected leukocytes in a fluorescent signals and the measurement was performed in a sheathless microfluidic channel thus minimizing the reagent volume to 68 ul in total, and consuming only a minimal blood sample of 5 ul. The whole system was enclosed in a suitcase sized package and the four-part differential count from this assay was validated by the results from test performed in central clinical laboratories.

The researchers concluded that the close correlation of this assay result with that of clinical laboratories along with the following merits:

  • minimal blood sample volume,
  • reagent long shelf life at room temperature,
  • lower reagent cost,
  • higher signal intensities,
  • portability

indicates that this method is ideal for a point-of-care assay. The researchers ultimate goal is to develop a portable device that can help patients living with chronic diseases such as leukemia at home. "It would be nice to limit the number of trips they need to make to the hospital for testing" says Wendian Shi [2], a graduate student in Tai's lab and lead author of the paper.

References

[1] Shi, W., et al., A Four-part leukocyte differential count based on sheathless microflow cytometer and fluorescent dye assay. Lab Chip, 2013, 13, 1257 DOI: 10.1039/c3lc41059e

[2] Counting White Blood Cells at Home. Science Daily http://www.sciencedaily.com/releases/2013/03/130326101608.htm 

Topics: Independent validation, Innovation, leukocyte differential, point of care test, research, white blood cells, whole blood

Maria

Written by Maria

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