There seems to be some consensus on the reaction mix for lactate dehydrogenase (LDH) activity assays, but the composition of the homogenization buffer (HB) differs wildly among studies. Maybe somebody can help me with the following three questions:

• Imidazole in HB: used in ~50% of studies in concentrations of 50-150 mM. Alternatively TRIS-HCl is used (50-75 mM). Almost all studies use imidazole in the reaction mix. Are there benefits to using imidazole in the HB instead of TRIS-HCl?
• Protease inhibitors (e.g., PMSF) in HB: used in few studies. Is this just for the workgroups that have more money to spend on chemicals?
• Some studies use KCN in the reaction mix without any explanation (earliest study that I could find was Sidell et al. 1987). Why?

Any recommendations and/or tips regarding LDH assays are greatly appreciated. Thanks!

Studies polled for this question:

Chippari-Gomes AR, Gomes LC, Lopes NP, Val AL, Almeida-Val VMF (2005) Comp Biochem Physiol B Biochem Mol Biol 141:347–355. https://doi.org/10.1016/j.cbpc.2005.04.006

Driedzic WR, Fonseca de Almeida-Val VM (1996) J. Exp. Zool. 274:327–333. https://doi.org/10.1002/(SICI)1097-010X(19960415)274:63.0.CO;2-Q

Gerringer ME, Yancey PH, Tikhonova OV, Vavilov NE, Zgoda VG, Davydov DR (2020) FEBS J 287:5394–5410. https://doi.org/10.1111/febs.15317

Heinrichs-Caldas W, Almeida-Val VMF de (2021) Fish Physiol Biochem 47:1759–1775. https://doi.org/10.1007/s10695-021-01000-0

Martinez ML, Chapman LJ, Grady JM, Rees BB (2004) J Fish Biol 65:1056–1069

Moon TW, Mommsen TP (1987) J. Exp. Zool. 244:9–15. https://doi.org/10.1002/jez.1402440103

Sidell BD, Driedzic WR, Stowe DB, Johnston IA (1987) Physiol Zool 60:221–232. https://doi.org/10.1086/physzool.60.2.30158646

Singer TD, Ballantyne JS (1989) J. Exp. Zool. 251:355–360. https://doi.org/10.1002/jez.1402510312