A whole-blood-staining device provides a means of (1) staining white blood cells by use of monoclonal antibodies conjugated to various fluorochromes, followed by (2) lysing and fixing of the cells by use of a commercial reagent that has been diluted according to NASA safety standards. More stable than whole blood, the lysed/fixed cells can be refrigerated at a temperature of 4 °C for as long as 72 h before processing and analysis on a flow cytometer.
This whole-blood-staining device is inexpensive and easy to manufacture. It offers advantages of compactness, robustness, and simplicity in comparison with equipment developed previously for the same purpose. The device is hand-held and self-contained. The use of the device does not require electric power, precise mixing, or precise incubation times.
The device (see figure) includes a reagent tube and two clips that separate the reagent tube into three compartments. During manufacture, the three compartments are loaded with (1) a solution containing the staining antibodies, (2) a lysing/fixing solution, and (3) a buffered saline solution, respectively. The first section, which contains the antibody solution, is equipped with a septum through which the blood sample can be injected.
At the time and place of sampling, 100 µL of anticoagulated whole blood is injected into the antibody solution via the septum. The device is shaken gently for about 10 s to mix the blood cells with the staining antibodies. The device is then held at room temperature for 30 min to incubate the blood-cell/antibody mixture. Next, the clip that separates the blood-cell/antibody mixture from the lysing/fixing solution is removed, the device is again shaken for about 10 s to mix the contents, and the device is again held at room temperature for 30 min. Then the other clip is removed and the device is shaken for about 10 s to mix in the saline solution. At this point, the device is refrigerated to preserve the mixture until further processing.
The device was developed for use in ascertaining changes in the immune systems of astronauts during long space flights. It is also suitable for use on Earth in small facilities that cannot afford expensive automated lysing equipment, large stocks of monoclonal-antibody reagents that could expire before use, and flow cytometers. Reference laboratories could send the devices to small rural facilities or doctors' offices for staining and lysing of blood samples, and the devices could be returned with the samples for analysis.
This work was done by Clarence F. Sams of Johnson Space Center; Vaughan Clift and Kelly E. McDonald of Martin Marietta Services, Inc.; and Ellen Meinelt of Krug Life Sciences.
This invention is owned by NASA, and a patent application has been filed. Inquiries concerning nonexclusive or exclusive license for its commercial development should be addressed to
the Patent Counsel
Johnson Space Center
Refer to MSC-22614