Figure 1

Isolation of crypts from a mouse colon. (A) Schematic of crypt isolation. The resected colon was incubated in chelating buffer, washed and then mechanically agitated. (B) Schematic of the strategy to identify the optimal acceleration intensity needed to retrieve crypts: the tissue was incubated in chelating buffer, rinsed and then sequentially agitated at different acceleration intensities. After each agitation, the crypt-rich supernatant was collected and assayed. (C) Fresh crypts isolated using an acceleration intensity of 1.5 × g displaying eGFP fluorescence, indicating Sox9 expression (green). Scale bar = 40 μm. (D) Quantification of the number of intact and broken crypts at each of the six acceleration intensities tested. Grey bars indicate intact-crypt yield and black bars indicate broken-crypt yield. (E) Brightfield images of isolated crypts isolated at different accelerations intensities. Crypts isolated using an acceleration intensity of 1.5 × g display a visible lumen, indicating unperturbed crypt morphology. Scale bar = 150 μm.