High-sensitivity flow cytometry provides access to standardized measurement of small-size microparticles--brief report.
OBJECTIVE
Cellular microparticles (MP) are promising biomarkers in many pathological situations. Although flow cytometry (FCM) is widely used for their measurement, it has raised controversies because the smallest MP size falls below the detection limit of standard FCM (sd-FCM). Following recent technological improvements leading to high sensitivity FCM (hs-FCM), our objectives were (1) to evaluate the potential of hs-FCM for extended MP detection, (2) to set up a standardized protocol for MP enumeration, and (3) to compare MP counts obtained with both sensitivity levels.


RESULTS
Compared with sd-FCM, hs-FCM displayed improved forward scatter resolution and lower background noise, allowing us to discriminate previously undetectable small MP in plasma samples. Using fluorescent beads with appropriate sizes (0.1/0.3/0.5/0.9 μm) and relative amounts, a new standardized hs-FCM MP protocol was set up and provided reproducible MP counts. Applied to coronary patient samples, it resulted into 8- to 20-fold increases in MP counts as compared with sd-FCM. Interestingly, the ratio between small and large MP varied according to clinical status but also depending on MP subset, suggesting access to new biological information.


CONCLUSIONS
Recent improvements in FCM provide access to previously undetectable MP and represent a new opportunity to enhance their impact as biomarkers in clinical practice.
UMR Institut National de la Santé et de la Recherche Médicale/Aix-Marseille Université, Faculté de Pharmacie, France.
Stéphane
S
Robert
Romaric
R
Lacroix
Philippe
P
Poncelet
Karim
K
Harhouri
Tarik
T
Bouriche
Coralie
C
Judicone
Jennifer
J
Wischhusen
Laurent
L
Arnaud
Françoise
F
Dignat-George
D015415:Biomarkers / Q000097:blood; D002138:Calibration; D055252:Cell-Derived Microparticles / Q000737:chemistry / Q000473:pathology*; D003327:Coronary Disease / Q000097:blood / Q000473:pathology*; D005434:Flow Cytometry / Q000379:methods / Q000592:standards; D006801:Humans; D010316:Particle Size; D011237:Predictive Value of Tests; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D015415:Biomarkers