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Clotspin basket

Manufactured by Qiagen
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The Clotspin Baskets are a specialized laboratory equipment used for the separation and isolation of clots or other solid components from liquid samples. The baskets provide a simple and efficient way to filter and collect these solid materials for further analysis or processing.

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Lab products found in correlation

Pathogen lysis tube, by Qiagen (4 mentions) Qiaamp ultraclean production pathogen mini kits, by Qiagen (4 mentions) Gentra puregene blood kit, by Qiagen (4 mentions) Nanodrop nd 1000 spectrophotometer, by Thermo Fisher Scientific (3 mentions) Proteinase k, by Qiagen (2 mentions) Rbc lysis solution, by Qiagen (1 mentions) Cell lysis solution, by Qiagen (1 mentions) Gentra systems autopure ls, by Qiagen (1 mentions) Masterpure complete dna rna purification kit, by Illumina (1 mentions) Hiscansq system, by Illumina (1 mentions) Genomestudio, by Illumina (1 mentions) Infinium humanmethylation450 beadchip, by Illumina (1 mentions) Geneamp pcr system 9700, by Thermo Fisher Scientific (1 mentions) Ez dna methylation kit, by Zymo Research (1 mentions) Atl buffer, by Qiagen (1 mentions) Buffer al, by Qiagen (1 mentions) Qiasymphony dsp dna midi kit, by Qiagen (1 mentions) Glycogen solution, by Qiagen (1 mentions) Picogreen dsdna quantitation kit, by Thermo Fisher Scientific (1 mentions) Oragene saliva collection kit, by DNA Genotek (1 mentions)

11 protocols using clotspin basket

1

DNA Isolation from Breast Milk and Saliva

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DNA was isolated from 1.5 mL of breast milk from each mother and from one Salivette saliva collection swab from each child. Before DNA isolation, Salivette cotton swabs were transferred to ClotSpin Baskets (Qiagen, Valencia, CA), thawed at room temperature, and centrifuged for 3 minutes at 2095 ×g to withdraw saliva into the ClotSpin Basket reservoir. Recovered saliva volumes ranged from 0.2 mL to over 1.5 mL. For those samples with greater than 1.5 mL recovered volume, only 1.5 mL was used for DNA isolation. Samples were subjected to a mechanical “bead beating” pretreatment using Pathogen Lysis Tubes (Qiagen, Valencia, CA) to disrupt Gram-positive bacterial cell walls according to the manufacturer’s protocol. Fat from breast milk samples was removed along with liquid supernatant after pelleting by centrifugation and before bead beating. DNA isolation from mechanically-lysed breast milk and saliva samples was completed using QIAamp Ultraclean Production Pathogen Mini Kits (Qiagen, Valencia, CA) according to the manufacturer’s protocol.
Davé V., Street K., Francis S., Bradman A., Riley L., Eskenazi B, & Holland N. (2016). Bacterial microbiome of breast milk and child saliva from low-income Mexican-American women and children. Pediatric research, 79(6), 846-854.
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2

DNA Isolation from Breast Milk and Saliva

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DNA was isolated from 1.5 mL of breast milk from each mother and from one Salivette saliva collection swab from each child. Before DNA isolation, Salivette cotton swabs were transferred to ClotSpin Baskets (Qiagen, Valencia, CA), thawed at room temperature, and centrifuged for 3 minutes at 2095 ×g to withdraw saliva into the ClotSpin Basket reservoir. Recovered saliva volumes ranged from 0.2 mL to over 1.5 mL. For those samples with greater than 1.5 mL recovered volume, only 1.5 mL was used for DNA isolation. Samples were subjected to a mechanical “bead beating” pretreatment using Pathogen Lysis Tubes (Qiagen, Valencia, CA) to disrupt Gram-positive bacterial cell walls according to the manufacturer’s protocol. Fat from breast milk samples was removed along with liquid supernatant after pelleting by centrifugation and before bead beating. DNA isolation from mechanically-lysed breast milk and saliva samples was completed using QIAamp Ultraclean Production Pathogen Mini Kits (Qiagen, Valencia, CA) according to the manufacturer’s protocol.
Davé V., Street K., Francis S., Bradman A., Riley L., Eskenazi B, & Holland N. (2016). Bacterial microbiome of breast milk and child saliva from low-income Mexican-American women and children. Pediatric research, 79(6), 846-854.
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3

Extracting High-Quality gDNA from Frozen Hematoma

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Frozen human fracture hematoma samples (n = 8) were thawed in a 37°C water bath and further kept on ice. To dissolve the hematoma samples, Clotspin® Baskets from Qiagen were used according the manufacturing protocol: Purification of archive-quality gDNA from clotted whole blood using Clotspin® Baskets and the Gentra® Puregene® blood kit provided by Qiagen. All protocol steps were followed according to the manufacturer's instructions. Steps included the usage of isopropanol and Glycogen Solution (20 mg/ml from Qiagen) and washing of gDNA pellets with 70% ethanol. gDNA was further air dried at room temperature until no remaining fluids remained. gDNA was incubated with 500 μl DNA hydration solution provided within the kit (www.qiagen.com/literature/handbooks/default.aspx). gDNA was incubated at 65°C until it was dissolved.
gDNA quality and quantity were confirmed using the NanoDrop-ND-1000 system (PEQLAB GmbH). Samples were stored at −80°C until further experiments.
Schlundt C., Reinke S., Geissler S., Bucher C.H., Giannini C., Märdian S., Dahne M., Kleber C., Samans B., Baron U., Duda G.N., Volk H.D, & Schmidt-Bleek K. (2019). Individual Effector/Regulator T Cell Ratios Impact Bone Regeneration. Frontiers in Immunology, 10, 1954.
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4

Genomic DNA Extraction from Cord Blood

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Genomic DNA from whole cord blood cells was extracted by the HiBR laboratory. Clotted cord blood was centrifuged through a Clotspin Basket (Qiagen) at 2,000 × g for 5 min to disperse the clot. Three volumes of Red Blood Cell (RBC) Lysis Solution (Qiagen) were added to one volume of dispersed cord blood. The samples were briefly vortexed, shaken for 15 min at room temperature then centrifuged as above to pellet leukocytes and clot particulates. The pellet was resuspended in Cell Lysis Solution (Qiagen) containing Proteinase K (Qiagen) at 20 mg/ml and then incubated at 55°C overnight. The Gentra Systems Autopure LS (Qiagen) was used to extract the DNA from the sample using the Compromised Cell Lysate Protocol. The purified DNA solution was then incubated in a 65°C water bath for 1 h and shaken at room temperature overnight.
Ching T., Ha J., Song M.A., Tiirikainen M., Molnar J., Berry M.J., Towner D, & Garmire L.X. (2015). Genome-scale hypomethylation in the cord blood DNAs associated with early onset preeclampsia. Clinical Epigenetics, 7(1), 21.
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5

DNA Methylation Analysis of Clotted Blood

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Total DNA was isolated from clotted blood samples using a clotspin basket (Qiagen, Hilden, Germany) to disperse the clot followed by extraction using the MasterPure Complete DNA & RNA Purification Kit (Epicentre, cat#MC85200). After the extraction, DNA concentration and purity (OD260/280) were measured using the NanoDrop ND-1000 spectrophotometer (Thermo Fischer Scientific, Waltham, MA, USA). A minimum of 500ng DNA was used for bisulfite conversion using the EZ DNA Methylation kit (Zymo Research Corp (Irvine, CA, USA) and a GeneAmp PCR system 9700 (Applied Biosystems, Grand Island, NY, USA). To confirm successful bisulfite modification, the DNA was subjected to PCR using methylation-specific primers. Methylation analysis was performed using the Illumina Infinium Human Methylation450 Beadchip platform (Illumina Inc., San Diego, CA, USA). Bisulfite-modified DNA was fragmented into 300–600 bp fragments, purified by isopropanol precipitation, and resuspended in hybridization buffer. The sample was then hybridized to an Illumina Infinium HumanMethylation450 Beadchip. The BeadChips were then washed, stained and dried. The BeadChips were then scanned using a HiScanSQ System (Illumina Inc., San Diego, CA, USA). Methylation data were processed through Illumina Genome Studio (Illumina Inc., San Diego, CA, USA) and analyzed in Partek.
Chen Y., Li X., Kobayashi I., Tsao D, & Mellman T.A. (2016). Expression and Methylation in Posttraumatic Stress Disorder and Resilience; Evidence of a role for Odorant Receptors. Psychiatry research, 245, 36-44.
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6

DNA Extraction from Blood Clots

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DNA was extracted from either blood clots remaining after serum separation by centrifugation or whole blood collected in EDTA tubes. In brief, clots were dispersed by centrifugation through clot spin baskets (Qiagen, 158932). ATL buffer (Qiagen, 1014758) was added to the centrifuged clot and vortexed. Proteinase K (Qiagen, 19131) was added, vortexed thoroughly and incubated in a shaking incubator at 56 °C until the clot was completely lysed (overnight). Afterr lysis, AL buffer was added (Qiagen, 1038826) and vortexed thoroughly. Lysate or whole blood was then transferred to the QIAsymphony 2.0 and extracted using the QIAsymphony DSP DNA Midi Kit (Qiagen, 937255).
Mentzer A.J., O’Connor D., Bibi S., Chelysheva I., Clutterbuck E.A., Demissie T., Dinesh T., Edwards N.J., Felle S., Feng S., Flaxman A.L., Karp-Tatham E., Li G., Liu X., Marchevsky N., Godfrey L., Makinson R., Bull M.B., Fowler J., Alamad B., Malinauskas T., Chong A.Y., Sanders K., Shaw R.H., Voysey M., Snape M.D., Pollard A.J., Lambe T, & Knight J.C. (2022). Human leukocyte antigen alleles associate with COVID-19 vaccine immunogenicity and risk of breakthrough infection. Nature Medicine, 29(1), 147-157.
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7

Whole Blood DNA Extraction Protocol

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DNA was isolated from clotted whole blood by using the Clotspin Baskets and the Gentra Puregene Blood Kit (Qiagen) according to the manufacturer’s instructions.
Watkin L.B., Jessen B., Wiszniewski W., Vece T., Jan M., Sha Y., Thamsen M., Santos-Cortez R.L., Lee K., Gambin T., Forbes L., Law C.S., Stray-Petersen A., Cheng M.H., Mace E.M., Anderson M.S., Liu D., Tang L.F., Nicholas S.K., Nahmod K., Makedonas G., Canter D., Kwok P.Y., Hicks J., Jones K.D., Penney S., Jhangiani S.N., Rosenblum M.D., Dell S.D., Waterfield M.R., Papa F.R., Muzny D.M., Zaitlen N., Leal S.M., Gonzaga-Jauregui C., Boerwinkle E., Eissa N.T., Gibbs R.A., Lupski J.R., Orange J.S, & Shum A.K. (2015). COPA mutations impair ER-Golgi transport causing hereditary autoimmune-mediated lung disease and arthritis. Nature genetics, 47(6), 654-660.
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8

DNA Extraction from Blood and Saliva

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DNA was isolated from blood clots using the Gentra Puregene Blood Kit combined with Clotspin Baskets (Qiagen, Hilden, Germany) or saliva using Oragene™ saliva collection kits according to manufacturer's instructions (DNA Genotek Inc., Ontario, Canada). All DNA samples were quantified using PicoGreen dsDNA quantitation kits (Molecular Probes, Eugene, OR).
de la Rosa R., Steinmaus C., Akers N.K., Conde L., Ferreccio C., Kalman D., Zhang K.R., Skibola C.F., Smith A.H., Zhang L, & Smith M.T. (2017). Associations between arsenic (+3 oxidation state) methyltransferase (AS3MT) and N-6 adenine-specific DNA methyltransferase 1 (N6AMT1) polymorphisms, arsenic metabolism, and cancer risk in a Chilean population. Environmental and molecular mutagenesis, 58(6), 411-422.
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9

DNA Extraction and Methylation Profiling

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For ACHS-I, DNA was extracted from stored blood clot samples using the PureGene (QIAGEN, 158,389) protocol and Clotspin Baskets (QIAGEN, 158,932). For ACHS-II, whole blood samples (300 µL per extraction) were aliquoted into deep well 96-well plates and genomic DNA was isolated using the Agencourt Genfind v2 Solid Phase Reversible Immobilization (SPRI) paramagnetic bead-based technology (Beckman Coulter, A41497). Prior to bisulphite-conversion, we measured DNA concentrations using a QUBIT dsDNA BR assay kit (Invitrogen, Q32850). The National Cancer Institute’s Cancer Genomics Research Laboratory performed DNA bisulphite conversion using the EZ-96 DNA Methylation MagPrep kit (Zymo Research, D5040) and then ran the bisulphite-converted DNA on Illumina EPIC methylation arrays (Illumina, WG-317-1001).
Pittman G.S., Wang X., Campbell M.R., Coulter S.J., Olson J.R., Pavuk M., Birnbaum L.S, & Bell D.A. (2019). Polychlorinated biphenyl exposure and DNA methylation in the Anniston Community Health Survey. Epigenetics, 15(4), 337-357.
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10

Blood and Saliva Collection for DNA Extraction

Cited 1 time
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The collection of specimens has been described previously [32 (link)]. Briefly, cases and controls provided a blood sample and completed a brief questionnaire at the time of blood draw, which updated breast and reproductive and gynecologic history and several lifestyle factors. Women who declined providing blood provided saliva in Oragene DNA self-collection kits (DNA Genotek, Kanata, ON, Canada). All biological specimens were sent overnight to the UCI laboratory. DNA was extracted from blood clots using Qiagen Clotspin Baskets and DNA QIAmp DNA Blood Maxi Kits (Qiagen, Inc., Valencia, CA, USA) in accordance with Qiagen protocols. DNA was extracted from saliva samples using the Oragene protocol (DNA Genotek).
Park H.L., Ziogas A., Chang J., Desai B., Bessonova L., Garner C., Lee E., Neuhausen S.L., Wang S.S., Ma H., Clague J., Reynolds P., Lacey JV J.r., Bernstein L, & Anton-Culver H. (2016). Novel polymorphisms in caspase-8 are associated with breast cancer risk in the California Teachers Study. BMC Cancer, 16, 14.
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