Optimal Cutting Temperature Medium‐Embedding Is a Valid Method for Storing and Preparing Myocardial Biopsies Preceding Myofilament Function‐Assessment

2020 ◽  
Vol 34 (S1) ◽  
pp. 1-1
Author(s):  
Henrik Isackson ◽  
Henry Ng ◽  
Mediha Becirovic Agic ◽  
Michael Hultström
Keyword(s):  
Cutting Temperature ◽  
Valid Method ◽  
Myofilament Function ◽  
Optimal Cutting Temperature ◽  
Myocardial Biopsies ◽  
Function Assessment

scholarly journals Optimal cutting temperature medium embedding and cryostat sectioning are valid for cardiac myofilament function assessment

2020 ◽  
Vol 319 (1) ◽  
pp. H235-H241
Author(s):  
Henry Ng ◽  
Mediha Becirovic Agic ◽  
Michael Hultström ◽  
Henrik Isackson
Keyword(s):  
Function Analysis ◽  
Cutting Temperature ◽  
Myocardial Tissue ◽  
Actomyosin Interaction ◽  
Myofilament Function ◽  
Passive Compliance ◽  
Cardiac Myofilament ◽  
Optimal Cutting Temperature ◽  
Force Activation ◽  
Function Assessment

Myocardial tissue in optimal cutting temperature (OCT) fixation and cryostat sectioning was tested as a means of storing and preparing tissue for myofilament function analysis in relation to conventional liquid nitrogen freezing and dissection. Actomyosin interaction, Ca2+ force activation, and passive compliance were tested. The study concluded that OCT storage and cryostat sectioning do not interfere with the actomyosin cross-bridge dynamics or Ca2+ activation but that absolute tension values suffer and may not be investigated by this method.

scholarly journals Cryo-Gel embedding compound for renal biopsy biobanking

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Malou L. H. Snijders ◽  
Marina Zajec ◽  
Laurens A. J. Walter ◽  
Remco M. A. A. de Louw ◽  
Monique H. A. Oomen ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Cutting Temperature ◽  
Renal Tissue ◽  
Tissue Samples ◽  
Normal Spleen ◽  
Renal Biopsies ◽  
Optimal Cutting Temperature ◽  
Embedding Medium ◽  
Rna And Dna

Abstract Optimal preservation and biobanking of renal tissue is vital for good diagnostics and subsequent research. Optimal cutting temperature (OCT) compound is a commonly used embedding medium for freezing tissue samples. However, due to interfering polymers in OCT, analysis as mass spectrometry (MS) is difficult. We investigated if the replacement of OCT with Cryo-Gel as embedding compound for renal biopsies would enable proteomics and not disturb other common techniques used in tissue diagnostics and research. For the present study, fresh renal samples were snap-frozen using Cryo-Gel, OCT and without embedding compound and evaluated using different techniques. In addition, tissue samples from normal spleen, skin, liver and colon were analyzed. Cryo-Gel embedded tissues showed good morphological preservation and no interference in immunohistochemical or immunofluorescent investigations. The quality of extracted RNA and DNA was good. The number of proteins identified using MS was similar between Cryo-Gel embedded samples, samples without embedding compound and OCT embedded samples. However, polymers in the OCT disturbed the signal in the MS, while this was not observed in the Cryo-Gel embedded samples. We conclude that embedding of renal biopsies in Cryo-Gel is an excellent and preferable alternative for OCT compound for both diagnostic and research purposes, especially in those cases where proteomic analysis might be necessary.

scholarly journals A simple method for sphingolipid analysis of tissues embedded in optimal cutting temperature compound

2020 ◽  
Vol 61 (6) ◽  
pp. 953-967
Author(s):  
Timothy D. Rohrbach ◽  
April E. Boyd ◽  
Pamela J. Grizzard ◽  
Sarah Spiegel ◽  
Jeremy Allegood ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cutting Temperature ◽  
Protein Quantification ◽  
Sphingolipid Metabolism ◽  
Normal Lung ◽  
Human Tissues ◽  
Simple Method ◽  
Disease Etiology ◽  
Esi Ms ◽  
Optimal Cutting Temperature

MS-assisted lipidomic tissue analysis is a valuable tool to assess sphingolipid metabolism dysfunction in disease. These analyses can reveal potential pharmacological targets or direct mechanistic studies to better understand the molecular underpinnings and influence of sphingolipid metabolism alterations on disease etiology. But procuring sufficient human tissues for adequately powered studies can be challenging. Therefore, biorepositories, which hold large collections of cryopreserved human tissues, are an ideal retrospective source of specimens. However, this resource has been vastly underutilized by lipid biologists, as the components of OCT compound used in cryopreservation are incompatible with MS analyses. Here, we report results indicating that OCT compound also interferes with protein quantification assays, and that the presence of OCT compound impacts the quantification of extracted sphingolipids by LC-ESI-MS/MS. We developed and validated a simple and inexpensive method that removes OCT compound from OCT compound-embedded tissues. Our results indicate that removal of OCT compound from cryopreserved tissues does not significantly affect the accuracy of sphingolipid measurements with LC-ESI-MS/MS. We used the validated method to analyze sphingolipid alterations in tumors compared with normal adjacent uninvolved lung tissues from individuals with lung cancer and to determine the long-term stability of sphingolipids in OCT compound-cryopreserved normal lung tissues. We show that lung cancer tumors have significantly altered sphingolipid profiles and that sphingolipids are stable for up to 16 years in OCT compound-cryopreserved normal lung tissues. This validated sphingolipidomic OCT compound-removal protocol should be a valuable addition to the lipid biologist’s toolbox.

scholarly journals Quantitative Proteomic Analysis of Optimal Cutting Temperature (OCT) Embedded Core-Needle Biopsy of Lung Cancer

2017 ◽  
Vol 28 (10) ◽  
pp. 2078-2089 ◽  
Author(s):  
Xiaozheng Zhao ◽  
Kenneth E. Huffman ◽  
Junya Fujimoto ◽  
Jamie Rodriguez Canales ◽  
Luc Girard ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Core Needle Biopsy ◽  
Proteomic Analysis ◽  
Needle Biopsy ◽  
Cutting Temperature ◽  
Core Needle ◽  
Quantitative Proteomic Analysis ◽  
Optimal Cutting Temperature

Depressed cardiac myofilament function in human diabetes mellitus

2005 ◽  
Vol 289 (6) ◽  
pp. H2478-H2483 ◽  
Author(s):  
Eias E. Jweied ◽  
Ronald D. McKinney ◽  
Lori A. Walker ◽  
Irwin Brodsky ◽  
Alexander S. Geha ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Artery Bypass ◽  
Cardiac Cells ◽  
Hill Coefficient ◽  
Diabetic Patients ◽  
Human Diabetes ◽  
Myofilament Function ◽  
Cardiac Myofilament ◽  
Separate Control ◽  
Myocardial Biopsies

Diabetes mellitus is associated with a distinct cardiomyopathy. Whether cardiac myofilament function is altered in human diabetes mellitus is unknown. Myocardial biopsies were obtained from seven diabetic patients and five control, nondiabetic patients undergoing coronary artery bypass surgery. Myofilament function was assessed by determination of the developed force-Ca2+ concentration relation in skinned cardiac cells from flash-frozen human biopsies. Separate control experiments revealed that flash freezing of biopsy specimens did not affect myofilament function. All patients in the diabetes mellitus cohort were classified as Type 2 diabetes mellitus patients, and most showed signs of diastolic dysfunction. Diabetes mellitus was associated with depressed myofilament function, that is, decreased Ca2+ sensitivity (29%, P < 0.05 vs. control) and a trend toward reduction of maximum Ca2+-saturated force (29%, P = 0.08 vs. control). The slope of the force-Ca2+ concentration relation (Hill coefficient) was not affected by diabetes, however. We conclude that human diabetes mellitus is associated with decreased cardiac myofilament function. Depressed cardiac myofilament Ca2+ responsiveness may underlie the decreased ventricular function characteristic of human diabetic cardiomyopathy.

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