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'''Protocol'''
*If testing fasted ketone tolerance: Remove food from mice for about 6h by putting them in a fresh cage. Add “do not feed” acetate to cages, and ideally move cage to procedure room. Try to make sure that the mice are in a quiet, undisturbed temperature controlled room with the lights on. **Typically starve the mice at 8AM and aim to start injections at 2PM
*Prepare a 1 g/10mL solution of glucose in the unlikely case that some animals become hyperketonemic.
*Weigh/MRI mice ahead of time, mark tails if necessary with different colors for rapid identification and take fasting ketone measurement via a tail clip.
*Prepare bOHB syringes with 1.5 g/KG mouse weight (ie for a 30g mouse, 300 uL).
*At approximately 1 min intervals, inject appropriate amount of bOHB into intraperitoneal cavity of the mouse. **Immobilize mouse and restrain tail with one hand. **Aim needle for peritoneal space, between the midline and the hip bone. **Insert syringe (do not inject) into cavity. **Eject syringe.
*At 15 minute intervals (normally 15, 30, 45, 60, 75, & 90 min), take blood bOHB measurements from tail vein. If needed re-snip the tail tip to expose the vein. When measuring BOHB just lift the tail of the mouse, while leaving it in the cage, rather than removing and restraining the mouse which can be more stressful.
*Analyze data by both % change from fasting ketones and absolute values. Our preferred outcome is to report fasting ketone levels and report percent change as a figure.
*The preferred statistical model is a mixed linear model using the time points as ordinate values and testing for a main effect or an interaction of the treatment/genotype. Use the R lme4 package for this.
[[Category:Ketogenesis]]