Bead Mill Homogenizers

Ocular tissues, ranging from small, soft perfused tissues to larger, tougher collagenous ones, are commonly selected for analysis during preclinical pharmacokinetic studies for ophthalmic drugs. The process of tissue homogenization and subsequent extraction needs to be optimized for proper downstream drug detection from these various matrices. In this study, bead mill homogenizers were selected for use for its efficiency and effectiveness in preparing complete homogenates. This methodology allows for the extraction of Cyclosporine from eight different ocular tissues – cornea, conjunctiva, iris-ciliary body (ICB), sclera, eyelid, lens, retina-choroid, and lacrimal gland. Due to the different phospholipid profiles between the tissues, however, varying amounts of matrix suppression were encountered during method development; to reduce the effect on analyte response, a phospholipid removal plate was used. The developed LC-MS/MS assay is robust, as well as demonstrates analytical reproducibility and efficiency for the quantification of Cyclosporine in the assayed ocular tissues. Acceptable selectivity, matrix factor, and dilution integrity were demonstrated in detail. Similarly, the methodology was flexible enough to allow for reanalysis of over-range samples as well as in posterior tissues with expected lower Cyclosporine levels. For research use only. Not for use in diagnostic procedures.

The success of sensitive downstream genomic applications such as qPCR depend on robust and reliable upstream sample preparation, which often includes proper sample/tissue homogenization and nucleic acid extraction. As the lab scales their sample throughput and their need for consistency in complex workflows, many have integrated automation-enabled protocols into their workflows. In this application note, a semi-automated workflow for nucleic acid extraction from liver tissue is presented, which leverages the Omni Bead Ruptor Elite bead mill homogenizer with the chemagic 360 nucleic acid extractor, with optional use of a JANUS automated liquid handling workstation.

Pathogenic fungi, i.e. dermatophytes, are responsible for superficial infections known as dermatophytosis, which can occur in nails, skin, and hair of humans and animals. In addition to dermatophytes, non-dermatophyte molds and yeasts can cause dermatomycosis. Though less of a risk in healthy individuals, the presence of these fungi can be severe in individuals with depleted or compromised immune systems. Therefore, research is being carried out to help optimize sample prep workflows and advance technologies that would help expedite species-specific pathogen identification. Aligned with this goal, this application note explores the efficiency of utilizing a semi-automated bead mill homogenizer in lieu of an overnight digestion in releasing nucleic acids from nail samples for downstream molecular genetic identification of dermatophytes, yeasts, and molds using the EUROArray dermatomycosis system. Results indicated good concordance between both research methodologies and significant time reduction in turn-around-time (TAT) when incorporating the bead mill homogenizer into the workflow as opposed to an overnight incubation.

Bead mill homogenizers enable rapid and efficient lysis of biological materials, including plants via high-impact forces generated from beads in an enclosed tube to dissociate samples. We explore the efficiency of vertical intra-tube bead milling motions of 7mL and 30mL tube and spiral grinding motions of a 50mL tube using the Omni Bead Ruptor Elite unit. Results indicate protein extraction yields were consistent run-to-run, in different tube sizes, and that full homogenization was achieved in less than a minute for all cases using the same homogenization settings within a given sample type. Thus, having one unit to support varying sample tube sizes allows bead milling to be leveraged for rapid and efficient sample homogenization across various sample input sizes/volumes for protein applications.

Bead mill homogenizers typically rely on vigorous shaking of tubes filled with bead material to disrupt samples. The impact between the beads and the sample reduces sample particle size and creates a homogenous mixture, making bead milling an efficient method of homogenization. Heat, however, is produced through the bead milling process because of the kinetic energy generated from the beads impacting the sample and tube walls. For some applications, such as those that are particularly heat-sensitive, labs could employ solutions to mitigate this issue as much as possible using cryo-cooling units. For some applications, the presence of the bead media within the sample may be unnecessary, such as for soft tissue homogenization. In this application note, we evaluated protein yields from rat brain tissue sections using “beadless” bead-milling homogenization on the Omni Bead Ruptor Elite bead mill homogenizer.

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This application note highlights the efficiency of bead mill homogenization on yeast cells, protected by a strong cell well. The Omni Bead Ruptor Elite bead mill homogenizer provided a rapid and efficient method for cell disruption to access cellular analytes in yeast models for molecular analysis. The process maintains the integrity of proteins for downstream molecular analyses like PAGE, Western blotting, and protein functional studies as noted by successful affinity capture chromatography.

Genomic and epigenomic information is important to scientific research, including human disease research. NGS (next-generation sequencing) is a popular technology for its ability to multiplex hundreds and thousands of samples for high-throughput workflows. To obtain meaningful data, however, researchers must depend on the initial quality and quantity of starting sample materials, which could be less than ideal in most cases. Labs, therefore, also depend on a reliable sample prep method upstream their preferred library prep assays. In this application note, we review a robust bead mill homogenization and multi-sample sonication method taken through ChIP-Seq analysis.

Sample prep is an important consideration for pre-clinical research utilizing animal models. Within sample prep, tissue dissociation and homogenization permits the analysis of analytes such as nucleic acids and proteins for downstream assays. Bead milling is an alternative to traditional disruption methods that may employ harsh chemicals or require numerous manual interventions under cryogenic conditions. In this application note, female nude mice were treated with an antimetabolic and common chemotherapeutic agent, 5-FU, which has been shown to have nephrotoxic and unwanted side effects following its use. Animal tissues were processed for total protein lysates and assayed for the biomarker phospho- and total-ERK, which has previously been found to be involved in acute and chronic kidney disease.

Bead mill sample homogenization, exemplified by the Omni Bead Ruptor Elite™ homogenizer, is a rapid and efficient method for sample disruption. However, for longer processing, the kinetic energy generated can increase sample temperature to potentially affect heat-sensitive analytes. We evaluated the passive cooling effect of the 7 mL carriage on larger volume samples, with the carriage and buffers pre-cooled to various temperatures. The results showed that using a carriage equilibrated at -80 °C in combination with chilled buffers effectively kept processing temperatures below 15 °C for up to 1 minute of processing time for larger volume samples though for samples that are in 2mL tubes, active chamber cooling is preferred utilizing a cryo cooling unit.

Bead mill technology is an efficient method for sample preparation in various fields. The kinetic energy, however, generated during the process can increase sample temperature, potentially affecting heat-sensitive analytes. The Omni BR-Cryo Cooling unit is designed to mitigate this issue by dissipating heat and maintaining lower intratube temperatures during the processing cycles on the Omni Bead Ruptor Elite. This complementary technology should be a key consideration when processing samples for the extraction of heat-sensitive molecules using bead milling.

We discuss a robust method of extracting bacterial DNA via bead milling for soil microbiome research. The Omni Bead Ruptor Elite bead mill homogenizer was used to lyse bacteria in soil samples for downstream 16S rRNA detection via endpoint PCR. The results demonstrate the homogenizer’s ability to extract microbial DNA from challenging soil samples such as Georgia red clay.

The comminution step in sample prep is critical for the effectiveness of the QuEChERS method, a type of dSPE, for the multiple pesticide extraction and detection. We evaluate the robustness of bead milling in 50 mL tubes for rapid homogenization of various types of certified organic produce, such as strawberries, soybeans, and apples spiked with 17 component pesticide standards at 100 ppb. Full homogenization by visual inspection was achieved in 30 seconds for all assayed samples of varying density with implemented settings. Pesticides were then extracted and analyzed by GC-MS for % recovery.

Nematodes, with their vast diversity, have adapted to various environments and include both parasitic and non-parasitic species. The vinegar eel, Turbatrix aceti , is a non-parasitic nematode used in DNA damage studies and is easy to maintain in unpasteurized vinegar. We demonstrate an efficient DNA extraction method from T. aceti using the Omni Bead Ruptor 4 bead mill homogenizer, which prepared high-integrity homogenates suitable for downstream DNA extraction. The optimizations help ensure minimal DNA shearing and consistent DNA concentration, highlighting the homogenizer’s effectiveness in sample preparation.

Drosophila melanogaster is a commonly used model organism in many research studies, including developmental biology. A reliable method for protein and nucleic acid extraction is a critical step toward extracting analytes that can help our understanding of molecular processes; this study demonstrates a fast and efficient method of extracting proteins and DNA from whole fruit flies using the Bead Ruptor Elite ™ bead mill homogenizer.

Hair is a widely studied sample matrix in various research fields, with DNA and metabolites being extracted for subsequent analysis. Traditional, manual methods of grinding hair can be inefficient and could potentially add risk of analyte loss or degradation due to time-spent grinding large sample sets. The Omni Bead Ruptor Elite bead mill homogenizer offers a solution, capable of grinding multiple hair samples in less than 10 minutes. This method results in a fine particle suspension or powder, suitable for downstream research analysis of hormones and toxicology metabolites.

Hair is a sample matrix that allows for non-invasive, fast and simple sampling. Small molecules including drug metabolites and hormones, proteins, and DNA can be extracted from hair samples from a wide variety of species. Extraction efficiency is low, however, so research laboratories often turn to automation-enabled bead mill homogenization to reduce particle size prior to carrying out further sample preparation procedures. In this application note, protocols on dry hair from various animal species are evaluated on the Omni Bead Ruptor Elite ™ bead mill homogenizer.

Milling is a crucial process in pharmaceutical production and process control, used to enhance solubility and stability of drug compounds through particle size reduction. We explored optimizing dry milling parameters to achieve effective particle size distributions, as dry milling is typically preferred for maintaining product integrity and scalability. On the Omni bead mill homogenizer, optimal milling frequency for analgesics ranged from 25 Hz and 30 Hz, with the most significant particle size reduction occurring within the first minute of milling. Extended milling times beyond five minutes showed negligible additional reduction.

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The efficiency of tissue homogenization is an important consideration upstream of many basic and pre-clinical research applications, including proteomic studies. Bead mill homogenization, such as with the Omni Bead Ruptor 12 homogenizer, facilitates uniform processing of multiple samples, achieving full homogenization in just 60 seconds to yield a consistent protein recovery of 7.2 ± 0.71 mg from ~50 mg of rat heart tissue, with less than 10% deviation as observed by SDS-PAGE. Labs can leverage the bead mill as an effective automation-enabled solution for high-throughput protein extraction from hard, dense tissues such as the heart in quantitative proteomic analyses compared to more manual methods.

The ability to detect pathogens throughout the food supply chain is an important component of food safety research. Just as the adoption of high-throughput methods such as PCR has expedited molecular detection of microorganisms vs more traditional time-consuming and labor-intensive methods, bead milling allows for an efficient and rapid sample prep method in foods, such as spinach. In this application note, both the bead mill and rotor-stator homogenizers were shown to fully process spinach samples spiked with recombinant E. coli while maintaining their viability. Both methods delivered a 76% cell recovery and a PCR detection limit of 1x10-6 CFUs/mL. Thus, the protocol and methods outlined may help expedite pathogen detection without necessitating a longer pre-enrichment step.

Hair samples are often used in forensic analysis research, including detection of performance-enhancing drugs like testosterone. Traditional methods of hair disruption for analysis have drawbacks, but bead milling, a method involving high-speed shaking of a sample with small beads, has been effective. The Omni Bead Ruptor Elite bead mill homogenizer, coupled with sonication, was used to extract free testosterone from equine hair samples. This method maintained the integrity of the testosterone, allowing for accurate measurement. The findings suggest bead milling is a promising technique for hair sample preparation in steroid or hormone analysis.