Figure 7.

Troubleshooting – examples of micropipette flow set incorrectly, resulting in loss of visualization

Incorrect balance pressure, such as too little (preventing just oozing beads and drug out of the micropipette), can cause an inflow from the bath liquid and tissue ECM, causing a loss of visualization of the micropipette tip (left image).

The blue arrowhead in both images points out the visible part of the micropipette that is present in a maximum intensity projection of the imaging stack.

The yellow arrowhead demonstrates where we thought the tip of the micropipette was when we started the run.

The purple arrowhead demonstrates where the tip actually was. With a loss of visualization, it is easy to wrongly center the z-stack in the tissue, resulting in the injection not being in frame and wasting the current image acquisition run.

Scale bars represent 50 μm.