24 h to 4 weeks post infection with M.tb, 3–7 mice per group were humanely euthanized with CO2. Lung and spleen tissue from infected animals were isolated and homogenized in 5 mL PBS + Tween-80 (Sigma-Aldrich, St. Louis, Missouri, USA) CFU buffer using an Omni tissue homogenizer (Omni International, Kennesaw, GA, USA). Tissue homogenates were serially diluted in CFU buffer and plated on Middlebrook 7H10 agar plates. Plates were incubated at 37 °C and with or without 5% CO2 for 2–3 weeks before colonies were counted. Bacterial burden, as CFU/mL, was calculated per organ and is presented as Log10 values.
Omni tissue homogenizer
Manufactured by Omni International
Sourced in United States
The Omni Tissue Homogenizer is a laboratory instrument designed to effectively disrupt and homogenize a wide range of biological samples, including tissues, cells, and other solid materials. The device utilizes high-speed rotation and advanced blade technology to efficiently break down samples into a homogeneous suspension, preparing them for further analysis or processing.
Automatically generated - may contain errors
Lab products found in correlation
Tween 80, by Merck Group (5 mentions)
Tri reagent, by Thermo Fisher Scientific (5 mentions)
In cell analyzer 6000, by GE Healthcare (3 mentions)
Rneasy mini kit, by Qiagen (3 mentions)
Protein assay, by Bio-Rad (2 mentions)
Protease inhibitor cocktail, by Roche (2 mentions)
Phosphotungstic acid pta, by Merck Group (2 mentions)
Rneasy minelute cleanup kit, by Qiagen (2 mentions)
Mirneasy mini kit, by Qiagen (2 mentions)
Nanodrop 2000c spectrophotometer, by Thermo Fisher Scientific (2 mentions)
Horse blood agar plates, by Thermo Fisher Scientific (2 mentions)
Hard tissue homogenizing tips, by Omni International (2 mentions)
Complete protease inhibitor, by Roche (2 mentions)
Ripa lysis buffer, by Thermo Fisher Scientific (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Edta free protease inhibitor cocktail, by Roche (1 mentions)
Quant ittm picogreentm dsdna assay kit, by Thermo Fisher Scientific (1 mentions)
Proteinase k, by Merck Group (1 mentions)
Waters 717 autosampler, by Waters Corporation (1 mentions)
515 hplc pump, by Waters Corporation (1 mentions)
60 protocols using omni tissue homogenizer
1
M.tb Infection Lung and Spleen CFU Quantification
2
Preparation of Human Alzheimer's Brain Extracts
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Brain tissues from 17 different human AD patients and 4 control individuals were obtained from brain banks affiliated with the University of British Columbia, the NIH NeuroBioBank and the National Prion Disease Pathology Surveillance Center (NPDPSC) at Case Western Reserve University. Donor characteristics are provided in Supplementary Table 1 . Informed consent for tissue collection at autopsy and neurodegenerative research use was obtained from the legal representative in accordance with local institutional review boards. These studies were reviewed and approved by the UBC Ethics Board and are in accordance with the Declaration of Helsinki principles. The clinical diagnosis of AD was based on NINCDS-ADRDA criteria. Samples from frontal cortex were weighed and submersed in ice-cold Tris-buffered saline (TBS) (20% w/v) with EDTA-free protease inhibitor cocktail (Roche Diagnostics, Laval QC, Canada), and homogenized using an Omni tissue homogenizer (Omni International Inc, Keenesaw GA, USA), 3 × 30 sec pulses with 30 sec pauses in between, all performed on ice. Homogenates were then subjected to ultracentrifugation at 100,000 × g for 60 min. Supernatants (soluble extracts) were collected, aliquoted and stored at −80 °C. The protein concentration was determined using a bicinchoninic acid (BCA) protein assay. Pools of brain extracts from 3–8 patients were used in each analysis.
3
Tissue Collection and RNA Extraction for VEEV
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Whole brain sections (frontal, parietal, temporal, occipital, cerebellum, thalamus), cervical lymph nodes (CLN), serum, cerebrospinal fluid (CSF), and nasal swabs were collected for VEEV control macaques. For whole brain sections and CLN, 100mg of tissue was harvested, suspended in 900 μL Tri-Reagent (Invitrogen), then homogenized using Omni tissue homogenizer (Omni International). For liquid samples, 100 μL of the specimen was suspended in 900μL Tri-Reagent, then thoroughly mixed by inversion. After a 10-minute incubation to ensure virus inactivation, the samples were transferred to a fresh tube prior to removal from the BSL-3 facility. Subsequent storage at -80°C or RNA isolation, cDNA synthesis, and qPCR analyses occurred in a Select Agent BSL-2 setting.
4
Quantification of Residual DNA in Decellularized Tissue
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Residual double stranded DNA (dsDNA) were quantified by fluorescent nuclei acid staining using the Quant-iTTM PicoGreenTM dsDNA Assay Kit (Molecular Probes, Inc., Eugene, OR). Samples were lyophilized and homogenized using a Omni Tissue Homogenizer (Omni, Kennesaw, GA) followed by incubation with 200 U/mL of Proteinase K (Sigma-Aldrich) for 16 h at 37 °C. Samples were then centrifuged at 2000 × g for 10 min and dsDNA quantified in the supernatant (ng dsDNA/initial wet weight of the sample) according to manufacturer’s instructions.
Fresh tissue submitted to detergent based decellularization33 (link) were used as positive controls. Briefly, pieces of PA were treated with a combination of 0.5% sodium deoxycholate (SDC) and 0.5% sodium dodecyl sulfate (SDS) for 24 h at room temperature with constant shaking. Samples were subsequently washed three times in phosphate buffered saline (PBS) for a total of 42 h, followed by fixation in formalin or quantification of residual DNA.
Fresh tissue submitted to detergent based decellularization33 (link) were used as positive controls. Briefly, pieces of PA were treated with a combination of 0.5% sodium deoxycholate (SDC) and 0.5% sodium dodecyl sulfate (SDS) for 24 h at room temperature with constant shaking. Samples were subsequently washed three times in phosphate buffered saline (PBS) for a total of 42 h, followed by fixation in formalin or quantification of residual DNA.
5
Quantification of α-Synuclein Prion Activity
6
Quantifying 5-HT and 5-HIAA in Mice
7
Cathepsin Enzyme Kinetics Assay
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The protocol for cathepsin enzyme kinetics was reported previously (34 (link)). In detail, GA muscles were dissected from wild type or Tpcn2−/− mice and extracted with 3 volumes of ice-cold cytosol lysis buffer (20 mm Tris-HCl, 1 mm EDTA, 1 mm EGTA, 1% glycerol, and freshly prepared dl -dithiothreitol (DTT) to a final concentration of 2 mm (pH 7.8)) with an OMNI tissue homogenizer (OMNI International). The pellets from centrifugation (13,000 × g, 30 min, 4 °C) were further extracted with 2 volumes of acid lysis buffer (200 mm sodium acetate, 50 mm NaCl, and 0.1% Triton X-100 (pH 5.0)), sonicated, and centrifuged again. The soluble lysates (15 μg) were then subjected to cathepsin enzyme kinetics assay with 20 μm fluorogenic peptide substrate Z-Phe-Arg-7-amino-4-methylcoumarin (Z-FR-AMC; Enzo Life Sciences) in cathepsin assay buffer (100 mm sodium acetate, 120 mm NaCl, and 1 mm EDTA (pH 5.5)). Duplicate lysates were preincubated with 50 μm E64d, a membrane-permeable cysteine protease inhibitor, for 15 min at 37 °C to inhibit cathepsin enzyme activities. Fluorescence readings derived from Z-FR-AMC hydrolysis were measured with a Flexstation 3 (Molecular Device) for 60 min at 37 °C. Fluorescence measurements were collected with 380-nm excitation and 460-nm emission at 1-min intervals. The acid enzyme kinetics (Vmax) was calculated as changes in relative fluorescence unit/min.
8
Viable Cell Counts of E. Coli Beads
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For viable cell counts, physical degradation of E. Coli bead samples was demonstrated using an Omni Tissue Homogenizer (Omni International). The samples were dissolved entirely by adding 100 μl of 0.1 M sodium citrate and back-diluted within 30 s in 1 ml of fresh LB media. The endpoint of a bacterial titer (10−7 to 20−10 dilutions) was then determined from the duplicate plates by standard plate count. 50 μl of the diluted samples were pipetted and dried on LB agar plates [38 (link)]. All samples on LB agar plates were incubated at 37 °C for 24 h. The average colony count between 30 and 150 colonies were considered for plate count.
9
Viable Cell Counts of E. Coli Beads
10
Western Blot Analysis of Mouse Liver Proteins
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Frozen mouse liver tissue was homogenized in RIPA buffer containing 1x EDTA-free protease inhibitor cocktail (Roche) using Omni Tissue Homogenizer (Omni International). The total protein concentration was determined by Bio-Rad Protein Assay and then equalized to 15 g/L. 25 μg of total protein was used for the western blot assay, performed as previously described (15 (link), 16 (link), 45 (link)). Antibodies used in the western blots are anti-BMAL1 (Cell signaling, #14020), anti-Wee1 (Cell signaling, #4936), and anti-TUBULIN (Sigma-Aldrich, T0198).
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