Fingernail Homogenization using a High Throughput Homogenizer
Keratin is a common protein of interest that is a major component of nails. Releasing keratin and other analytes from nails can be very difficult due to the very tough nature of the tissue. Nail samples can be ground cryogenically, but this is slow and can be a major bottleneck in sample preparation.
Keratin is a resilient structural protein resulting from the complex network formed by keratin monomers. It is the bundled keratin proteins and the layers of dead, flattened cells that gives fingernails and toenails their strength. However, fingernails can easily be homogenized using a high throughput homogenizer and a stainless steel grinding ball. A standard protocol for fingernail homogenization can be found below. The protocol was designed using human fingernails. Multiple samples may be homogenized using a 24 Well PC Vial Set.
Weigh 20 to 40 mg of nail clippings and place into a 4 ml polycarbonate vial. For very tough tissues like nail, it is very important to use polycarbonate vials due to their hard and rigid design. Vials made of polyethylene or polypropylene tend to soften as they warm during sample processing and this can lead to less effective homogenization.
Insert a 3/8" stainless steel grinding ball and securely fasten the lid. As no extraction buffer is used in this protocol, the cap used for closing the vial can be either lined or unlined.
Place the vials into a high through put homogenizer, such as an HT Homogenizer or GenoGrinder. Vials can be placed in racks or in a vial sets (up to 24 to 48 samples per run, depending on the homogenizer). Homogenize at 1500 rpm for five minutes. Remove the vials and examine the contents. If small pieces of nail remain, repeat the process.
Keratin is very resilient and difficult to homogenize. Traditional homogenizers, such as rotor-stators, are ineffective at processing nails, and manual grinding methods are laborious. A bead mill such as the HT Homogenizer and GenoGrinder provide sufficient power when using large grinding balls to effectively pulverize nails.
Once ground, the nails will appear as a fine powder, though when mixed with an extraction buffer, the relatively insoluble keratin will appear as small flecks. Microscopic observation (such as the picture to the left) demonstrates that the nail has been reduced signficantly.