Zinc buffered formalin fixative

Liver fibrosis was induced in mice using the hepatotoxic agent carbon tetrachloride (CCl₄) (cat# 270652, Sigma-Aldrich, St. Louis, MO, USA), as previously described [43 (link)]. Briefly, 4-week-old male C57BL/6J mice nourished with normal chow and water were given intraperitoneal (IP) injections of 0.2 μL/g CCl₄ twice weekly for 20 weeks. During each IP injection, mice were briefly anesthetized in an induction chamber using a 2% isofluorane/O₂ mixture to minimize animal movement and the risk of needle sticks. The CCl₄ was administered as an 8% CCl₄/corn oil solution. Therefore, for a typical 20 g mouse, 50 μL of the solution was administered with each IP injection. A fresh 8% CCl₄/corn oil solution was created each week. Control mice were given identical IP injections of pure corn oil. After 20 weeks, mice were anesthetized, sacrificed by cervical dislocation, and the livers weighed and removed. Livers were dissected into equal halves, one of which was flash frozen in liquid nitrogen and stored at -80°C for later molecular analysis. The other half was suspended in zinc-buffered formalin fixative (cat# 5701ZF, Thermo Fisher Scientific, Waltham, MA, USA) for three days before it could be mounted in a paraffin block for histological analysis.

Liver fibrosis was induced in mice using the hepatotoxic agent carbon tetrachloride (CCl₄) (cat# 270652, Sigma-Aldrich, St. Louis, MO, USA), as previously described [24 (link)]. Briefly, 10-week-old male C57BL/6J mice nourished on normal chow and water were given intraperitoneal (IP) injections of 0.2 uL/g CCl₄ twice weekly for 20 weeks. During each IP injection, mice were briefly anesthetized in an induction chamber using a 2% isofluorane/O₂ mixture to minimize animal movement and the risk of needle sticks. The CCl₄ was administered as an 8% CCl₄/corn oil solution. Therefore, for a typical 20 g mouse, 50 uL of the solution was administered with each IP injection. A fresh 8% CCl₄/corn oil solution was created each week. Control mice were given identical IP injections of pure corn oil. After 20 weeks, mice were anesthetized, sacrificed by cervical dislocation, and the livers weighed and removed. Livers were dissected into equal halves, one of which was flash frozen in liquid nitrogen and stored at -80°C for later molecular analysis. The other half was suspended in zinc-buffered formalin fixative (cat# 5701ZF, Thermo Fisher Scientific, Waltham, MA, USA) for three days before it could be mounted in a paraffin block for histological analysis.