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HelpOrder Code DHRF Dihydrorhodamine Flow Cytometric N-Formyl-Methionyl-Leucyl-Phenylalanine Test, Blood
Useful For
Evaluation of Rac2 deficiency and RAC2 gain of function
Reporting Name
DHR Flow fMLP, BSpecimen Type
WB Sodium HeparinShipping Instructions
Testing performed Monday through Friday. Specimens not received by 4 p.m. Central time on Fridays may be canceled.
Collect and package specimen as close to shipping time as possible. Ship specimen overnight in an Ambient Shipping Box-Critical Specimens Only (T668) following the instructions in the mailer. It is recommended that specimens arrive within 24 hours of collection.
Specimens arriving on the weekend and observed holidays may be canceled.
Necessary Information
Ordering healthcare professional name and phone number are required.
Specimen Required
Two whole-blood sodium heparin specimens are required, one from the testing patient and the other from an unrelated healthy donor as a control.
Supplies: Ambient Shipping Box-Critical Specimens Only (T668)
Patient:
Container/Tube: Green top (sodium heparin)
Specimen Volume: 5 mL
Collection Instructions: Send whole blood specimen in original tube. Do not aliquot.
Normal Control:
Container/Tube: Green top (sodium heparin)
Specimen Volume: 5 mL
Collection Instructions:
1. Collect a control specimen from the unrelated healthy donor within an hour of the patient's specimen collection time.
2. Clearly label as Normal Control on the outermost label.
3. Send the whole blood specimen in the original tube. Do not aliquot.
4. Rubber band patient specimen and control vial together.
Specimen Minimum Volume
1 mL
Specimen Stability Information
| Specimen Type | Temperature | Time | Special Container |
|---|---|---|---|
| WB Sodium Heparin | Ambient | 48 hours | GREEN TOP/HEP |
Reject Due To
| Gross hemolysis | Reject |
| Gross lipemia | Reject |
Reference Values
|
Result name |
Unit |
Cutoff for defining normal |
|
% fMLP ox-DHR+ |
% |
≥10% |
|
MFI fMLP ox-DHR+ |
MFI |
≥2 |
|
Control % fMLP ox-DHR+ |
% |
≥10% |
|
Control MFI fMLP ox-DHR+ |
MFI |
≥2 |
fMLP = N-formyl-methionyl-leucyl-phenylalanine
DHR = dihydrorhodamine
MFI = mean fluorescence intensity
The appropriate age-related reference values for Absolute Neutrophil Count will be provided on the report.
Method Description
A sodium heparin whole blood specimen is incubated at 37° C in the presence of DHR123. N-formyl-methionyl-leucyl-phenylalanine (fMLP) stimulant is added and mixed with the whole blood specimen for additional incubation at 37° C. The sample is then centrifuged, and cell pellet is subsequently lysed with ammonium chloride at room temperature. Lysed samples are then washed with azide-free phosphate buffered saline prior to staining with LIVE/DEAD viability marker and CD15 at ambient temperature. Finally, cells are washed, centrifuged, and resuspended in 1% paraformaldehyde prior to analysis. Viable neutrophils are identified by the use of the viability dye and further confirmed by the presence of CD15. Approximately 20,000 viable neutrophil events in the unstimulated sample are used to set the limits for number of events collected for flow cytometry. The results are derived as delta % DHR+ neutrophils after fMLP stimulation and mean fluorescence intensity.(O'Gorman MR, Corrochano V. Rapid whole-blood flow cytometry assay for diagnosis of chronic granulomatous disease. Clin Diagn Lab Immunol. 1995;2[2]:227-232; Kuhns DB. Diagnostic testing for chronic granulomatous disease. Methods Mol Biol. 2019;1982:543-571)
Performing Laboratory
Mayo Clinic Laboratories in Rochester
CPT Code Information
86352
Method Name
Flow Cytometry
Day(s) Performed
Monday through Friday