Multiple In Vivo Liver Biopsies Using a Freeze-Clamping Technique

Abstract

A multiple in vivo liver biopsy technique was developed to measure labile metabolites (creative phosphate [CP], ATP, lactate) without interfering with normal perfusion and metabolism. Anesthetized rats (n = 7) had a midline abdominal incision done to expose the liver. Four biopsies were taken across 20 min. Bleeding was controlled by a small, nontraumatic clamp proximal to the biopsy. Prefrozen dressing forceps grasped the liver and scissors cut the biopsy, which was frozen in liquid nitrogen. Bleeding was minimal and hemodynamic stability was preserved. This technique has few complications, bleeding is easily controlled, and it allows for large multiple biopsies, which give reliable metabolic data and can be consistently taught.

Impact of Female Sex Hormones on Liver Tissue Lactic Acidosis During Ischemia

“…2 and 3) (28). All groups displayed similar significant increases in lactate by 5 min of ischemia (PϽ0.001).…”

“…To profile the time course of lactate and H ϩ accumulation, full thickness biopsies were taken at 5, 10, 15, 30, and 45 min of normothermic ischemia, which includes time points that are seen clinically (9) as well as time points (45 min) beyond the recommended safe limits (30 min) (13). Biopsies were taken using the flash-freeze clamp technique (28) and stored at Ϫ85°Celsius until analysis for lactate and H ϩ .…”

“…A carotid artery catheter was used to monitor blood pressure, heart rate, and to sample blood to ensure normal blood gases. A midline incision was performed to expose the liver, and an in vivo biopsy was taken using the flash-freeze clamp technique (28). A large section of the liver was removed from its blood supply (onset of ischemia) and maintained normothermic by a Krebs-Henseleit bath for the duration of the ischemic study.…”

Diet-induced mouse model of fatty liver disease and nonalcoholic steatohepatitis reflecting clinical disease progression and methods of assessment

“…Furthermore, the ability to sample hepatic tissue from the live animal facilitates studies to identify objective diagnostic and prognostic markers of disease obtained by less invasive techniques. Methods incorporating clinical biopsy guns, diathermy, or freeze-clamping to control blood loss in rats (3,7,8) have not been widely adopted. To reduce the risk of tissue injury that can result from biopsy, we chose Gelfoam absorbable gelatin sponge to control bleeding (17).…”

Promises and pitfalls of untargeted metabolomics

“…A variety of extraction/quenching methodologies have been compared for tissue that has been excised or biopsied from animals. Issues that have warranted extensive investigation include the need to cryo-freeze tissue, the use of freeze clamping, as well as variables associated with animal anesthesia and euthanasia methods (Belanger et al 2002; Want et al 2013; Overmyer et al 2015). In addition, the extraction solution used can have a major influence on the scope of the metabolome observed.…”