title: Rapid detection of circulating fibrocytes by flowcytometry in idiopathic pulmonary fibrosis.
abstract: BACKGROUND
  Current protocols for detection of circulating fibrocytes (CFs) in peripheral blood described in various pulmonary and nonpulmonary disorders involve complex and time consuming, non standardized techniques.
  
  
  OBJECTIVE
  Testing a method to rapidly detect and quantify CFs using whole blood lysis flow cytometry-based assay in patients with idiopathic pulmonary fibrosis (IPF) and healthy controls.
  
  
  METHODS
  One milliliter of venous blood sample in ethylenediaminetetraacetic acid (EDTA) from 33 IPF patients and 35 healthy control subjects was collected. Using whole blood lysis method peripheral blood leukocytes were labeled with monoclonal antibodies for cell surface (CD34 and CD45) and intracellular markers (collagen-1) for flow cytometric analysis. CFs were defined as CD45(+) cells coexpressing collagen-I and CD34 molecules.
  
  
  RESULTS
  In 29 (87.8%) IPF patients and 10 (28.5%) control subjects, a well-defined highly granular CD45(+) cell population was detected in dot plots generated by side scatter properties of CD45(+) cells. These CD45(+) cells were identified as CFs on the basis of coexpression of collagen-I and CD34; none of the other cell types in the peripheral blood were labeled with these monoclonal antibodies. In IPF patients the percentage of CFs was significantly higher compared to healthy controls (median (range): 1.37% (0.52-5.65) and 1.04% (0.1-1.84), respectively; P = 0.03).
  
  
  CONCLUSIONS
  Whole blood lysis method combined with fluorescence-activated cell sorting (FACS) allows detecting a well-defined homogeneous population of CFs. This method is simple, reproducible, and provides an accurate and rapid estimation of CFs.
authors.affiliation: Department of Medicine, King Khalid University Hospital, College of Medicine, King Saud University, Riyadh, Saudi Arabia.
authors.forename: Esam H
authors.initials: EH
authors.lastname: Alhamad
authors.affiliation: Department of Pathology, King Khalid University Hospital, College of Medicine, King Saud University, Riyadh, Saudi Arabia.
authors.forename: Zahid
authors.initials: Z
authors.lastname: Shakoor
authors.affiliation: Department of Medicine, King Khalid University Hospital, College of Medicine, King Saud University, Riyadh, Saudi Arabia.
authors.forename: Feisal A
authors.initials: FA
authors.lastname: Al-Kassimi
authors.affiliation: Department of Pathology, King Khalid University Hospital, College of Medicine, King Saud University, Riyadh, Saudi Arabia.
authors.forename: Adel
authors.initials: A
authors.lastname: Almogren
authors.affiliation: Department of Pathology, King Khalid University Hospital, College of Medicine, King Saud University, Riyadh, Saudi Arabia.
authors.forename: Mohamed O
authors.initials: MO
authors.lastname: Gad ElRab
authors.affiliation: Firestone Institute for Respiratory Health, Department of Medicine, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Shyam
authors.initials: S
authors.lastname: Maharaj
authors.affiliation: Firestone Institute for Respiratory Health, Department of Medicine, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Martin
authors.initials: M
authors.lastname: Kolb
keywords: Bonemarrow-derived cells; circulating fibrocytes; fibrosis; flow cytometry; idiopathic pulmonary fibrosis