title: Validation of Sysmex XT-2000iV generated quantitative bone marrow differential counts in untreated Wistar rats.
abstract: BACKGROUND
  Preclinical drug trials frequently require assessment of bone marrow toxicity in animals to evaluate hematopoietic safety. Since the gold standard, cytologic evaluation, is time consuming and requires highly trained individuals, automated methods remain intriguing.
  
  
  OBJECTIVE
  The Sysmex XT-2000iV hematology analyzer allows user-developed customizable gating. This study was conducted to validate the gating of bone marrow cell populations in Sysmex cytograms from untreated rats.
  
  
  METHODS
  B- and T-lymphocytes and myeloid cells were experimentally depleted from Charles River Wistar Han IGS (CRL: WI [Han]) rat whole bone marrow suspension using a magnetic cell sorting (MACS) method. The positively and negatively selected populations were used to verify select gates within the Sysmex cytogram. Intra- and inter-animal precision, comparability between right and left femur, as well as agreement with microscopic myelograms based on 500 counted cells, were determined.
  
  
  RESULTS
  Intra-sample precision and right-to-left femur comparability confirmed that gating was reproducible and stable. In 50 tested rats, myeloid to erythroid ratios (M:E) were 1.32 ± 0.33 in males and 1.38 ± 0.29 in females by Sysmex compared to 1.36 ± 0.32 in males and 1.42 ± 0.32 in females by microscopic evaluations. Bland-Altman differences between methods was ≤ ± 0.35 units for M:E, ≤ 5.4% for maturing myeloid cells, ≤ 3.4% for proliferating myeloid cells, ≤ 6.0% for maturing myeloid cells, ≤ 3.4% for proliferating myeloid cells, and ≤ 4.1% for lymphocytes.
  
  
  CONCLUSIONS
  In untreated control Charles River Wistar Han IGS (CRL: WI [Han]) rats, the Sysmex XT-2000iV produced an automated M:E and 5-part differential count equivalent to microscopic differential counts.
authors.affiliation: Pfizer Worldwide Research and Development, Drug Safety Research and Development, Groton, CT, USA.
authors.forename: Kay A
authors.initials: KA
authors.lastname: Criswell
authors.forename: Jeffrey H
authors.initials: JH
authors.lastname: Bock
authors.forename: Samantha E
authors.initials: SE
authors.lastname: Wildeboer
authors.forename: Kjell
authors.initials: K
authors.lastname: Johnson
authors.forename: Richard P
authors.initials: RP
authors.lastname: Giovanelli
mesh_terms: D000818:Animals; D001322:Autoanalysis / Q000295:instrumentation / Q000662:veterinary*; D001854:Bone Marrow Cells / Q000166:cytology*; D001856:Bone Marrow Examination / Q000662:veterinary; D002452:Cell Count / Q000662:veterinary; D049109:Cell Proliferation; D002470:Cell Survival; D004353:Drug Evaluation, Preclinical / Q000662:veterinary; D041905:Erythroid Cells / Q000166:cytology; D005260:Female; D005434:Flow Cytometry / Q000295:instrumentation / Q000662:veterinary; D008214:Lymphocytes / Q000166:cytology; D008297:Male; D022423:Myeloid Cells / Q000166:cytology; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results
keywords: Flow cytometry; hematotoxicity; myelotoxicity; preclinical development; preclinical safety