The DBS punching was performed using a Wallac DBS Puncher (PerkinElmer, Waltham, MA, USA) into wells of 96-well plates. The 3.2 mm diameter punches underwent semi-automated DNA extraction in a Tecan Robotic Liquid Handler (Tecan, US, Morrisville, NC, USA) using Generation DNA Purification and Elution Solutions (QIAGEN, Germantown, MD, USA). The extraction protocol included two purification washes of the DBS by adding 100 µL of Generation DNA Purification solution per well, followed by one wash with Generation DNA Elution Solution. All washes were performed at 37 °C for 10 min on a microplate shaker. After discarding the Elution buffer, 75 µL of new Elution solution was added to each well and incubated for 30 min at 98 °C while being shaken in a VorTemp 56 (Labnet, Edison, NJ, USA) at 1300 rpm. Plates were then cooled down to ambient temperature.
Wallac dbs puncher
Manufactured by PerkinElmer
Sourced in United States
The Wallac DBS Puncher is a compact, automated instrument designed for extracting samples from dried blood spot (DBS) cards. It is capable of precisely punching out circular samples from DBS cards, which can then be used for various analytical tests and applications.
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Lab products found in correlation
5 protocols using wallac dbs puncher
1
Automated DNA Extraction from DBS Samples
2
Quantifying Amino Acids and Acylcarnitines
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Each sample was analyzed for 12 amino acids and 31 acylcarnitines [a 3.2 mm circle was obtained using a Wallac DBS Puncher (PerkinElmer, Waltham, MA, USA)] from the dry filter paper. A NeoBase non-derivatized LC-MS/MS kit (Perkin Elmer) was used to obtain the metabolites of interest, following the manufacturer’s instructions. A solution included in the kit containing internal standards labeled with stable isotopes was used for quantifying the metabolites of interest. The samples were analyzed by LC-MS/MS (API 2000, ABSciex, Framingham, MA, USA) coupled to a micropump and an autosampler (Series 2000, Perkin Elmer). Sample analysis was performed with multiple reaction monitoring using Analyst 1.6.2 Software (ABSciex) and the NeoBase database. The results were interpreted using Analyst 1.6.2 Software.
3
Screening for Sickle Cell Disease
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Screening for SCD was performed at MUHAS HCRL: 3.2 mm of the DBS was punched by an automated DBS puncher (Wallac DBS Puncher) from PerkinElmer, Massachusetts, USA followed by IEF analysis, conducted according to the manufacturer’s protocol (Wallac RESOLVE Haemoglobin System). Samples showing abnormal results such as those found with the sickle gene (FAS), those found to be positive for SCD (FS) and those found to have unidentified variants FAV for the first test were re-punched and retested by HPLC. Results were interpreted by a trained laboratory technician and confirmed by the laboratory manager. Confirmed results were entered into the program’s database using the unique sample identification number.
4
Dried Blood Spot Sample Preparation
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Whatman 903TM paper cards were purchased from Sigma Aldrich (St. Louis, MO, USA). DMPD cartridges (disk diameter; 5 mm) and 10 µL fixed-volume capillary micropipettes were provided by Rock Town Technologies & Services (Libertyville, IL, USA). 5 mm paper discs were obtained using a WALLAC DBS Puncher (Perkin Elmer, Waltham, MA, USA).
5
DNA Extraction from Saliva and Newborn Spots
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Saliva samples were collected using the Oragene® DNA Self-Collection Kit (DNA Genotek, Ottawa, Canada) from both twins, their PM mother and maternal aunt (Figure 1 ). A single saliva swab (DNA Genotek, Global) collected orally per participant, was processed as per manufacturer’s instructions. DNA was extracted using QIAsymphony DSP DNA extraction Kit (Qiagen, Hilden, Germany) at the Victorian Clinical Genetics Services (VCGS), with DNA quality and concentration assessed by NanoDrop 2000 spectrophotometer. Newborn Blood Spots (NBS) from the MZ twins were retrieved from the VCGS NBS repositories as part of the FREE FX study. One 3-mm punch from each NBS disk was placed in a 1.5-mL Eppendorf tube using the Wallac DBS Puncher (Perkin Elmer, Waltham, MA, USA), and incubated in 55 μL in salt lysis buffer (Murdoch Children’s Research Institute, Melbourne, Australia) for 15 min in a heat block for cell lysis, degradation of proteins and release of the DNA, as previously described [13 (link)].
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