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Automated immunostaining of cell smears: an alternative to flow cytometry
  1. L C Happerfield1,
  2. R Saward2,
  3. L Grimwade2,
  4. D Bloxham2,
  5. W N Erber2
  1. 1
    Histopathology Department, Cambridge University Hospitals NHS Foundation Trust (Addenbrooke’s Hospital), Cambridge, UK
  2. 2
    Haematology Department, Cambridge University Hospitals NHS Foundation Trust (Addenbrooke’s Hospital), Cambridge, UK
  1. Dr W Erber, Haematology Department (Box 234), Addenbrooke’s Hospital, Cambridge University Hospitals NHS Foundation Trust, Hills Road, Cambridge CB2 0QQ, UK; wendy.erber{at}


Aims: To assess the applicability of an automated tissue immunostainer machine for the phenotypic analysis of peripheral blood and bone marrow aspirate smears.

Methods: Air-dried smears of peripheral blood and bone marrow from normal individuals and 14 patients with haematological malignancies were stained, following fixation, with a range of antibodies to haemopoietic antigens using both immunoperoxidase and immuno-alkaline phosphatase methods on the Bond-maX (Leica Microsystems) automated immunostaining machine.

Results: Automated protocols used for staining formalin-fixed paraffin-embedded tissue could be applied to cell smears, giving high quality staining with excellent cytological detail and antigenic preservation for an extensive antibody panel. Optimal quality (morphological preservation and antigen detection without background staining) was obtained with an alkaline phosphatase method and Fast Red chromogenic substrate. Both cell surface and intracellular (cytoplasmic and nuclear) antigens could be detected and gave the expected reactivity.

Conclusions: Automated immunostaining is possible for haematological smears. It generates consistent high quality staining, with the exception of surface immunoglobulin, and is applicable to haematological and, potentially, cytological smears. It provides a practical alternative to flow cytometry and will have particular application when smears are available and there is no suitable sample for flow cytometry.

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  • Competing interests: None.