title: A comparison of manual and automated methods of quantitation of oestrogen/progesterone receptor expression in breast carcinoma.
abstract: BACKGROUND
  Oestrogen/progesterone receptor expression in breast carcinoma is associated with good response to hormonal therapy and overall better prognosis. The predictive and prognostic capabilities of these receptors are enhanced by quantitation of immunoreaction. There are several manual and automated methods for this purpose. Whether they yield comparable results that can be used interchangeably is not yet clear.
  
  
  OBJECTIVE
  To compare the manual methods (H-score and Allred score) with automated methods (Immunoratio) for quantifying immunohistochemical (IHC) reaction for ER/PR in breast carcinoma.
  
  
  METHODS
  Samples from established cases of breast carcinoma were processed and stained by immunohistochemical methods to demonstrate oestrogen receptor (ER) and progesterone receptor (PR). Receptor expression was quantified by manual methods (H-score, modified H-score and Allred score) and automated methods (basic and advanced Immunoratio). In modified H score, the intensity of reaction was assessed by measurement of mean grey value {H (MGV)} or optical density {H (DC-OD)} of deconvoluted image. The manual counting was done with cell counter plugin of Image-J (NIH). The scores were compared and Pearson's correlation coefficient was determined.
  
  
  RESULTS
  Both manual and automated methods produced results that were comparable. There was a statistically significant positive correlation among all methods (p<0.02). The strongest correlation was observed between advanced immunoratio and H (DC-OD) (p=<0.001). Basic immunoratio appeared to be less reliable than the other methods. Staining intensity measurements by various methods did not significantly affect correlation. However, intensity measurements by optical density resulted in lower H-scores but led to more reliable detection of negative immunoreaction.
  
  
  CONCLUSIONS
  Both manual and automated methods of quantitation are comparable. Advanced immunoratio is a reliable alternative to manual methods. Cell Counter plugin is a useful tool for manual counting and quantitation.
authors.affiliation: Associate Professor, Department of Pathology, Chettinad Hospital & Research Institute , Kelambakkam, Tamilnadu, India .
authors.forename: R
authors.initials: R
authors.lastname: Vijayashree
authors.affiliation: Post-graduate Department of Pathology, Chettinad Hospital & Research Institute , Kelambakkam, Tamilnadu, India .
authors.forename: P
authors.initials: P
authors.lastname: Aruthra
authors.affiliation: Professor & HOD, Department of Pathology, Chettinad Hospital & Research Institute , Kelambakkam, Tamilnadu, India .
authors.forename: K Ramesh
authors.initials: KR
authors.lastname: Rao
keywords: Allred score; Cell counter plugin; Deconvolution; ER; Image-J; PR; immunoRatio; ‘H’ score