In diagnostic histopathology, hematoxylin and eosin (H&E) staining is a critical process that highlights salient histological features. Staining results vary between laboratories regardless of the histopathological task, although the method does not change. This variance can impair the accuracy of algorithms and histopathologists’ time-to-insight. Investigating this variance can help calibrate stain normalization tasks to reverse this negative potential. With machine learning, this study evaluated the staining variance between different laboratories on three tissue types. We received H&E-stained slides from 66 different laboratories. Each slide contained kidney, skin, and colon tissue samples stained by the method routinely used in each laboratory. The samples were digitized and summarized as red, green, and blue channel histograms. Dimensions were reduced using principal component analysis. The data projected by principal components were inserted into the k-means clustering algorithm and the k-nearest neighbors classifier with the laboratories as the target. The k-means silhouette index indicated that K = 2 clusters had the best separability in all tissue types. The supervised classification result showed laboratory effects and tissue-type bias. Both supervised and unsupervised approaches suggested that tissue type also affected inter-laboratory variance. We suggest tissue type to also be considered upon choosing the staining and color-normalization approach.

Recent developments in deep learning have impacted medical science. However, new privacy issues and regulatory frameworks have hindered medical data sharing and collection. Deep learning is a very data-intensive process for which such regulatory limitations limit the potential for new breakthroughs and collaborations. However, generating medically accurate synthetic data can alleviate privacy issues and potentially augment deep learning pipelines. This study presents generative adversarial neural networks capable of generating realistic images of knee joint X-rays with varying osteoarthritis severity. We offer 320,000 synthetic (DeepFake) X-ray images from training with 5,556 real images. We validated our models regarding medical accuracy with 15 medical experts and for augmentation effects with an osteoarthritis severity classification task. We devised a survey of 30 real and 30 DeepFake images for medical experts. The result showed that on average, more DeepFakes were mistaken for real than the reverse. The result signified sufficient DeepFake realism for deceiving the medical experts. Finally, our DeepFakes improved classification accuracy in an osteoarthritis severity classification task with scarce real data and transfer learning. In addition, in the same classification task, we replaced all real training data with DeepFakes and suffered only a $$3.79\%$$ 3.79 % loss from baseline accuracy in classifying real osteoarthritis X-rays.