Image analysis of liver collagen using sirius red is more accurate and correlates better with serum fibrosis markers than trichrome

Abstract Background Collagen proportional area ( CPA ) determined by quantitative digital image analysis better quantifies liver fibrosis than histological stage; however, its clinical use has been limited by non‐standardized methods. Aim This study aimed to compare CPA obtained using different staining methods, magnifications and biopsy sizes. Methods Two hundred and forty‐nine patients with chronic hepatitis C who had a liver biopsy and serum fibrosis markers performed were included. CPA was measured either using a sirius red ( CPA s) or a trichrome ( CPA t) stain. Results CPA s measured at 20× and 40× magnifications generated similar outcomes with interclass correlation ( ICC ) coefficient of 0.98. Compared with trichrome, sirius red staining had much less variation with an ICC coefficient of 0.99 for slides stained in the same batch and 0.92 in different batches. Mean CPA s was higher than mean CPA t by 3.53%, P < 0.001. Morphological analysis found that sirius red detected delicate fibrous septa and spurs better than trichrome. Both CPA s and CPA t correlated well with Metavir stage, whereas CPA s had better ability to detect cirrhosis with the area under ROC curve of 0.95. Overall CPA had superior correlation with serum markers of fibrosis in Metavir F2–F4 than that in F0–F1 and CPA s correlated better with serum fibrosis markers than CPA t in Metavir F0–F1. Multivairate analysis found that HA , α2‐macroglobulin, platelet count and albumin were independently correlated with CPA s and only HA was independently correlated with CPA t. Conclusions Sirius red staining for CPA determination was more accurate and reliable for quantifying hepatic collagen compared with trichrome staining.