High Throughput Grinding / Homogenization for DNA Extraction and Protein Isolation
BT&C/OPS Diagnostics
672 Rt. 202-206 North
Bridgewater, NJ
TEL: (908) 253-3444
FAX: (908) 575-1660
Email: info@btc-bti.com
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Maintenance of Protein Stability
High throughput DNA extraction and protein isolation often requires homogenization / grinding of hard samples, such as seeds, and large pieces of tissue (animal and plant). This can be complicated as the samples are not readily adaptable to methods using standard 96 deep well plates. For seeds, the size of and inherent toughness often require pre-treatment and/or dissection prior to adding them to deep well plates. Also, the standard 5/32 inch (5 mm) stainless steel grinding balls used in deep well plates do not have sufficient mass to fracture larger seeds, thus first soaking the seeds to soften them is often necessary. With corn, kernels are often halved after soaking so that they will fit into wells. Overall, these time consuming manipulations do not lend themselves to high throughput methodologies.
OPS Diagnostics has developed tools using the standard 24 well plate format to speed the high throughput homogenization process. The 24 Well Vial Set combines rugged polycarbonate vials, large 3/8 inch stainless steel grinding balls, and a racking system in a standard and familiar format. Each 24 Well Vial Set comes assembled with 24 vials, 24 balls, rack with foam insert, a sealing pad, and 24 polypropylene caps.
With 24 Well Vial Sets and a Geno/Grinder 2000 (or similar ball mill), 48 corn kernels can be pulverized into a dry powder in 2 minutes. This allows for the processing of approximately 1000 samples per hour, a rate suitable for even PCR analysis. Once homogenized, grinding balls are easily removed with the 24 Pin Magnet. A Magnet Stand/Separator has also been developed to aid in dropping balls into the vials and for separating balls from the 24 Pin Magnet. For laboratories with lower throughput, Magnet Tips can be used to remove balls from vials and deep well plates. These modified Pipetteman tips can be used alone or with a multichannel pipettor to remove 8 or 12 balls at a time.
For researchers who require pooled samples (as with field trials) for analysis, the 15 ml Vial Set is available. These larger vials can hold up to 15 corn kernels per grinding. Each vial contains two large stainless steel grinding balls and there are 5 vials per set.
For both the 24 Well Vial Sets and 15 ml Vial Sets, homogenized samples can be subsequently processed by standard RNA isolation, DNA extraction, or protein purification methods.
Method
The 24 Well Vial Set is a pre-assembled grinding media that includes a rack, vials, grinding balls, vial caps and sealing mat. The mat is removed and the balls are extracted manually or by using the 24 Pin Magnet. Samples are then placed in each well and the balls are replaced either manually or by using the Magnet Stand/ Separator. The lid of the box is removed and the sealing mat is placed over the vials (Fig. 1). The vial set and mat are then locked into the Geno/Grinder (or similar grinder). The locking fixture of the Geno/Grinder presses the sealing mat over and into the vials. The mat is not glued or heat sealed. The samples are then homogenized, and then removed from the grinder. The mat is carefully removed and the balls are extracted with the 24 Pin Magnet.
For protocols where RNA and DNA extraction will be performed, it is advised to dip the magnet in 3% bleach between samplings in order to reduce the chance of cross-contamination. The polypropylene caps can be used to seal individual vials if cross-contamination is a concern for subsequent PCR analysis. These vials can be removed and handled individually.
Homogenized seeds and plant tissue in HTS
format
Figure 1. 24 Well Vial Set complete with sealing mat. Each box contains vials, b
Figure 2. The grinding balls are easily removed with the 24 Pin Magnet before the adding the samples and replaced using the Magnet Stand/Separator.
Products and Accessories
(For a complete list of products see www.opsdiagnostics.com)
Specifically designed for high throughput homogenization of kernels and other seeds. Each set contains 24 polycarbonate tubes, 24 stainless steel 3/8 inch grinding balls, 24 vial caps, and one sealing mat. Vials can be capped and handled as individual samples following grinding.
24 Well Vial Sets are also available with polyethylene vials. These less expense PE Vial Sets are suitable for most small seeds and animal tissue. These are not recommended for corn kernels. Patent Pending.
24 Well PC Vial Set (polycarbonate vials) $240/case of 10 sets
24 Well PE Vial Set (polyethylene vials) $195/case of 10 sets
Components of a 24 Well Vial Set
The 24 Pin Magnet is used to efficiently remove grinding balls from the 24 Well Vial Set. When used with the Magnet Stand/Separator, the 24 Pin Magnet can serve to both remove and replace grinding balls into the vials. Patent Pending.
Price: $450
The Magnet Stand/Separator is used to align the 24 Well Vial Set and the 24 Pin Magnet to efficiently remove and/or replace the grinding balls into vials. The magnet fits into the stand which is positioned above the vial set. The magnet then slides downward with the pins descending into the wells to retrieve the balls. The balls are removed by lifting a slide. Alternatively, the slide drops the pins with balls into the vials and by lifting the magnet, the balls separate from the magnet and fall into the vials. The Magnet Stand/Separator is designed to be adaptable to motorization. Patent Pending.
Price: Call!
Magnet Tips are a simple, cost effective solution for removing stainless steel grinding balls from 96 well plates and grinding vials. The tips are made for Pipetteman micropipettes. For other pipettes, custom tips are available. Call for details. Patent Pending.
Pack of 12 Magnet Tips $58
Rack of 96 Magnet Tips $410
The 15 ml Vial Set was developed for researchers who required pooled samples for analysis. This product was designed for the difficult job of dry grinding 15 lyophilized corn kernels for field trials. Each 15 ml Vial Set comes with 5 polycarbonate vials per box with two stainless steel grinding balls per vial. Patent Pending.
Price for case of 10 sets, 5 vials per set (50 vials total): $240
For small grinding media, such as silica beads or zirconium beads, see the grinding media.