Effects of Different Storage Conditions on Lipid Stability in Mice Tissue Homogenates

Abstract

Lipids are biomolecules involved in numerous (patho-)physiological processes and their elucidation in tissue samples is of particular interest. However, tissue analysis goes hand in hand with many challenges and the influence of pre-analytical factors can intensively change lipid concentrations ex vivo, compromising the results of the whole research project. Here, we study the influence of pre-analytical factors on lipid profiles during the processing of homogenized tissues. Homogenates from four different mice tissues (liver, kidney, heart, spleen) were stored at room temperature as well as in ice water for up to 120 min and analyzed via ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS). Lipid class ratios were calculated since their suitability as indicators for sample stability has been previously illustrated. Only approx. 40% of lipid class ratios were unchanged after 35 min, which was further reduced to 25% after 120 min during storage at room temperature. In contrast, lipids in tissue homogenates were generally stable when samples were kept in ice water, as more than 90% of investigated lipid class ratios remained unchanged after 35 min. Ultimately, swift processing of tissue homogenates under cooled conditions represents a viable option for lipid analysis and pre-analytical factors require more attention to achieve reliable results.

Evaluation of Lipid Extraction Protocols for Untargeted Analysis of Mouse Tissue Lipidome

“…Several techniques have been used for tissue homogenization, such as grinding frozen tissue with mortar and pestle [ 37 ] or bead-beating-based methods [ 38 , 39 ]. Since homogenization has been evaluated and optimized elsewhere [ 27 , 40 ], in the present study, tissue homogenization was decoupled from the lipid extraction evaluation. The preparation of a fluidic aqueous homogenate of each tissue, followed by fast aliquoting of the homogenate and subsequent lipid extraction of each aliquot, was adopted in the present study.…”

Differential Lipidomics, Metabolomics and Immunological Analysis of Alcoholic and Non-Alcoholic Steatohepatitis in Mice

“…For extraction, liver homogenates were diluted using pre-cooled extraction solution to tissue concentrations of 0.05 mg/µL. Lipids and polar metabolites were extracted as previously described via liquid–liquid extraction using 20 µL of tissue homogenate, corresponding to 1 mg of tissue material [ 40 , 41 ]. Samples were kept in ice water during processing and the extracted and dried samples were stored at <−70 °C until analysis.…”

Study on the Properties and Synergistic Antioxidant Effects of Novel Bifunctional Fusion Proteins Expressed Using the UTuT6 System

“…By adding 20 mL of pre-cooled 0.9% saline per 1 g of liver, liver homogenate with a defined concentration (5% m / v ) was created by using a homogenizer. Protein amount was determined with Coomassie Brilliant Blue using BSA as the standard [ 23 ]. The incubation mixture consisted of 0.05 mM GSH, 5% liver homogenate, appropriate enzymes, and 100 mM H 2 O 2 .…”