Patent ID: 8428331

Claim:
A method of counting a cluster of unstained cells, comprising the steps of: (a) providing a microscope capable of first and second imaging modalities, wherein the first imaging modality is a full-field phase imaging technique and the second imaging modality provides cell boundary information for the unstained cell cluster; (b) obtaining a first image of the cell cluster, the image including biological material surrounding the cell cluster and a culture medium in which the cell cluster is suspended, using the first imaging modality and a second image of the cell cluster using the second imaging modality, wherein the first image is an image of optical path length deviation (OPD); (c) fitting an ellipse to the boundary of a cell in the second image; (d) determining the maximum optical path length deviation of the cell in the first image and determining the minimum optical path length deviation of the cell as the optical path length deviation of the biological material surrounding the cluster of cells plus the optical path length deviation of the culture medium in which the cell cluster is suspended in the first image, and calculating the total optical path length deviation of the cell as the maximum optical path length deviation of the cell minus the minimum optical path length deviation of the cell; (e) combining the ellipse with the total optical path length deviation to produce an ellipsoidal model cell of optical path length deviation; (f) subtracting the model cell from the OPD image to obtain a subtracted OPD image; and (g) repeating steps (c), (e), and (f), wherein for each repetition of step (f) the model cell is subtracted from the previous subtracted OPD image, until model cells corresponding to all cell boundaries detectable in the second image have been subtracted from the OPD image; wherein the cell count for the cell cluster is the total number of model cells subtracted in step (f).