The following tissues were disrupted at full speed for 30 seconds using the TissueRuptor: frozen kidney, liver, and spleen (10 mg each); frozen lung (250 mg); stabilized heart, muscle, and brain (10 mg each). Samples were either disrupted in lysis buffer (kidney, liver, spleen, lung, heart, and muscle) or QIAzol Lysis Reagent (brain). Total RNA was purified using the RNeasy Plus Mini Kit (RNeasy Plus), the RNeasy Midi Kit (RNeasy Midi), the RNeasy Fibrous Tissue Mini Kit (RNeasy Fibrous), or the RNeasy Lipid Tissue Mini Kit (RNeasy Lipid). Purified RNA was analyzed by formaldehyde agarose gel electrophoresis. The gels show the high yields and quality of the RNA following disruption using the TissueRuptor.
Frozen liver samples (30 mg each) were disrupted at full speed for 30 seconds using either the TissueRuptor with disposable probes or a traditional rotor–stator homogenizer with a steel generator probe. Total RNA was purified using the RNeasy Plus Mini Kit and analyzed on the Agilent 2100 Bioanalyzer. The high RNA Integrity Number (RIN) indicates the high quality of the RNA.
[A] Liver and [B] heart samples (25 mg each, frozen) were disrupted with the TissueRuptor or a traditional rotor–stator homogenizer and lysed for 1 hour with proteinase K, or lysed overnight with proteinase K without sample disruption. DNA was purified using the QIAamp DNA Mini Kit. Eluates (5 μl) were subjected to PCR using HotStarTaq DNA Polymerase and primers specific for the 18S ribosomal RNA gene. M: GelPilot Mid Range Ladder.
Frozen rat liver and brain (approximately 30 mg each) were disrupted at medium speed for 30–60 seconds in 350 μl PhosphoProtein Lysis Buffer. The homogenized samples were then diluted with 1500 μl PhosphoProtein Lysis Buffer and incubated at 4°C for 30 minutes, with brief vortexing every 10 minutes. Phosphoproteins were purified using the PhosphoProtein Purification Kit and detected by western blotting. [A] Transfer membrane stained with Ponceau S, and [B] western blot. M: markers; FT: flow-through; E3: elution fraction 3.
Frozen liver and kidney samples (30 mg each) were disrupted in lysis buffer at full speed for 30 seconds using the TissueRuptor. Genomic DNA was purified using the AllPrep DNA/RNA Mini Kit and analyzed by agarose gel electrophoresis. M: markers.