30 μm pre separation filter

Manufactured by Miltenyi Biotec

Sourced in Germany

The 30 μm pre-separation filter is a laboratory equipment used to remove large particles and cell aggregates from cell suspensions prior to cell isolation or other downstream applications. It is designed to fit standard cell preparation tubes and syringes, allowing for easy integration into various cell processing workflows.

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Lab products found in correlation

Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions) Ephb4, by R&D Systems (2 mentions) Ephrinb2, by R&D Systems (2 mentions) Rat igg2a, by R&D Systems (2 mentions) Facscalibur, by BD (2 mentions) Facsaria 2, by BD (2 mentions) 70 μm cell strainer, by BD (1 mentions) Til isolation kit, by Miltenyi Biotec (1 mentions) Gentlemacs octo dissociator, by Miltenyi Biotec (1 mentions) Whole blood column elution buffer, by Miltenyi Biotec (1 mentions) Quadromacs separator, by Miltenyi Biotec (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions) Anti cd45 magnetic beads conjugated antibody, by Miltenyi Biotec (1 mentions) Hydrated ms columns, by Miltenyi Biotec (1 mentions) Propidium iodide, by Beckman Coulter (1 mentions) Propidium iodide, by BD (1 mentions) Accutase, by Innovative Cell Technologies (1 mentions) Epics xl, by Beckman Coulter (1 mentions) Liberase solution, by Roche (1 mentions) Dnase 1, by New England Biolabs (1 mentions)

Close spelling variants of this product

Pre-Separation Filters (29 citations) 30 μm filter (12 citations) 70-μm and 30-μm pre-separation filters (2 citations)

11 protocols using 30 μm pre separation filter

Naïve T Cell Isolation from Mouse Spleen

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Lymphocytes suspension was prepared from fresh mouse spleens by gently grinding the spleens between two glass slides. The lymphocytes suspension was filtered through 70-μm cell strainers (BD Biosciences) to make single lymphocytes suspension. This lymphocytes suspension was further filtered through 30-μm Pre-Separation Filters (Miltenyi Biotec). Then, CD4⁺CD62L⁺ naïve T cells were isolated using mouse CD4⁺CD62L⁺ T Cell Isolation Kit II and MidiMACS™ Separator, following the manufacturer’s instructions. The collected cells were stained with anti-CD4-FITC and anti-CD62L-allophycocyanin antibodies (Miltenyi Biotec) and analyzed with flow cytometry, which showed that the isolated lymphocytes were = 92% positive for both CD4 and CD62L that were considered as naïve T cells (40 (link)).

Chen R.Y., Fan Y.M., Zhang Q., Liu S., Li Q., Ke G.L., Li C, & You Z. (2015). Estradiol Inhibits Th17 Cell Differentiation through Inhibition of RORγT Transcription by Recruiting ERα/REA Complex to the EREs of RORγT Promoter. Journal of immunology (Baltimore, Md. : 1950), 194(8), 4019-4028.

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Isolation and Analysis of Thymic and Splenic Immune Cells

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For ex vivo analysis of thymocytes and CD8⁺ T-cells derived from spleens and LNs of female 8–12-week old WT, LMP7-deficient and TKO mice were used. The single-cell suspensions were generated by mechanical disruption of thymi, spleens, and LNs using 30 μm pre-separation filters (Miltenyi Biotec). Afterwards, single-cell suspensions were counted before proceeding with the flow cytometry.

Leister H., Luu M., Staudenraus D., Krol A.L., Mollenkopf H.J., Sharma A., Schmerer N., Schulte L.N., Bertrams W., Schmeck B., Bosmann M., Steinhoff U, & Visekruna A. (2021). Pro- and anti-tumorigenic capacity of immunoproteasomes in shaping the tumor microenvironment. Cancer immunology research, 9(6), 682-692.

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Isolation and Characterization of Tumor-Infiltrating Lymphocytes in Melanoma

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B16-F10 melanoma cells were cultured in RPMI 1640 medium (Thermo Fisher Scientific) supplemented with 100 U/ml penicillin/streptomycin, 10% FCS and 2 mM glutamine for 1 week before inoculation into mice. WT, TKO LMP7^−/− mice were subcutaneously (s.c.) injected with 1 × 10⁶ B16-F10 cells. In some experiments, the B16-GFP cell line was used as well. Tumor growth was monitored daily for 15 days by caliper measurements. To analyze T cells in tumors and tumor-draining lymph nodes (LNs), mice were sacrificed, and tumor tissues and draining LNs were harvested. The single-cell suspensions from LNs were obtained by mechanical disruption using 30 μm pre-separation filters (Miltenyi Biotec). Tumor-infiltrating lymphocytes (TILs) were isolated using the Miltenyi TIL Isolation kit (Miltenyi Biotec, 130–096-730) and the GentleMACS Octo Dissociator (Miltenyi Biotec) according to the manufacturer’s instructions. Briefly, the melanoma tumors were cut into small pieces and the tissue was transferred into GentleMACS tubes containing the enzyme mix. The tumors were dissociated into single-cell suspensions by combining enzymatic digestion with mechanical dissociation. After termination of the program, the cell suspensions were centrifuged (300xg, 7 minutes) and supernatant was completely aspirated.

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EpCAM-Based Positive Selection for Cell Isolation

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A positive selection-based MACS technology was used (Miltenyi Biotec, Bergisch Gladbach, Germany) following the manufacturer’s instructions. The sample was first filtered through a 30 μm pre-separation filter (Miltenyi Biotec). The sample was then incubated with anti-EpCAM magnetic beads (StraightFrom^® Whole Blood CD326 (EpCAM) MicroBeads, Miltenyi Biotec, 50 μl/ml blood, 30 min, 2–8°C) for magnetic labeling, followed by centrifugation (445 rpm, 10 min) and resuspension of the cell fraction with a separation buffer. Magnetic separation was performed using a column containing a matrix composed of ferromagnetic spheres (Whole Blood Columns, Miltenyi Biotec) placed in a separator (QuadroMACS Separator, Miltenyi Biotec). This was followed by the elution step (Whole Blood Column Elution Buffer, Miltenyi Biotec, 5 ml), yielding an enriched, positively selected fraction. All collection tubes were extensively washed with separation buffer throughout the procedure in order to avoid any potential cell loss.

Lozar T., Jesenko T., Kloboves Prevodnik V., Cemazar M., Hosta V., Jericevic A., Nolde N, & Grasic Kuhar C. (2020). Preclinical and Clinical Evaluation of Magnetic-Activated Cell Separation Technology for CTC Isolation in Breast Cancer. Frontiers in Oncology, 10, 554554.

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Murine Splenocyte Isolation Protocol

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For splenocytes, the spleen from a C57BL/6Jax mouse was mashed by a 2-ml syringe plunger through a 70 μm cell strainer (Fisher Scientific 10788201) into 30 ml 1X DPBS (Thermo Fisher 14190169) supplied with 2 mM EDTA and 0.5% (w/v) BSA (Sigma A9418). Cells were centrifuged down, supernatant was removed, and the cell pellet was briefly vortexed. 5 ml 1X RBC lysis buffer (Thermo Fisher 00-4300-54) was used to resuspend the cell pellet, and the cell suspension was vortexed again, and left on bench for 5 min to lyse red blood cells. Then 45 ml 1X DPBS was added, and cells were centrifuged down. Volume of 30 ml 1X DPBS was used to resuspend the cell pellet. The cell suspension was passed through a Miltenyi 30 μm Pre-Separation Filter (Miltenyi 130-041-407), and the cell number was determined using C-chip counting chamber (VWR DHC-N01). All centrifugations were done at 500×g, 4 °C, 5 min. For human and mouse skin fibroblasts, cells were extracted as previously described¹⁵. For mouse cardiac progenitor cells, cells were extracted as previously described¹⁶. Cells were cryopreserved in 90% FBS and 10% DMSO and stored in liquid nitrogen until experiments.

Chen X., Miragaia R.J., Natarajan K.N, & Teichmann S.A. (2018). A rapid and robust method for single cell chromatin accessibility profiling. Nature Communications, 9, 5345.

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Isolation of Stromal Vascular Cells and Mitochondria

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Stromal vascular cells were resuspended in 500 mL of magnetic bead buffer (MBB) (PBS 1× without calcium and magnesium, 0.5% w/v BSA, and 2 μM ethylenediaminetetraacetic acid [EDTA]) and filtered through a 30-μM pre-separation filter (Miltenyi, Bergisch Gladbach, Germany) following three filter washes to remove large particles and debris. The cell suspension was separated at 300 × g for 5 min at 4°C and resuspended in 90 mL of MBB plus 10 mL of anti-CD45 magnetic beads-conjugated antibody (Miltenyi) for each 10⁷ cell sample. After 15 min of incubation at 4°C, the cell suspension was diluted with 2 mL of MBB and centrifuged. The cell pellet was resuspended in 500 mL of MBB, applied on hydrated MS-columns (Miltenyi), washed three times with 500 mL of MBB, and collected with 1 mL of MBB through piston elution. Incubation with anti-CD11b and anti-F4/80 magnetic beads-conjugated antibodies (Miltenyi) was performed in a unique incubation, as was performed for CD45. BAT was homogenized using a Potter–Elvehjem polytetrafluoroethylene pestle and a glass tube and TO μM 22⁺ mitochondria were magnetically isolated by a commercially available kit (Miltenyi).

Rosina M., Ceci V., Turchi R., Chuan L., Borcherding N., Sciarretta F., Sánchez-Díaz M., Tortolici F., Karlinsey K., Chiurchiù V., Fuoco C., Giwa R., Field R.L., Audano M., Arena S., Palma A., Riccio F., Shamsi F., Renzone G., Verri M., Crescenzi A., Rizza S., Faienza F., Filomeni G., Kooijman S., Rufini S., de Vries A.A., Scaloni A., Mitro N., Tseng Y.H., Hidalgo A., Zhou B., Brestoff J.R., Aquilano K, & Lettieri-Barbato D. (2022). Ejection of damaged mitochondria and their removal by macrophages ensure efficient thermogenesis in brown adipose tissue. Cell metabolism, 34(4), 533-548.e12.

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Analyzing Cell Cycle in NKX2-5-GFP hESCs

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Differentiated NKX2–5-GFP hESCs at different time points were dissociated with Accutase (Innovative Cell Technologies, San Diego, CA, USA) for 30 min, and passed through a 30 μm pre-separation filter (Miltenyi Biotec Inc., Auburn, CA, USA) to produce a single cell suspension. The cells were analyzed on EPICS XL (Beckman Coulter, Miami, FL, USA) with Expo32 ADC XL 4 Color software. Data analysis was performed using FlowJo Software (Version X; TreeStar, Ashland, OR, USA). Cell sorting was performed using Fluorescence Activated Cell Sorting (FACS) on EPICS XL (Beckman Coulter).
NKX2–5-GFP ES cells were treated with MG-132 at specific concentrations (25 μM) and with UA (7.5 mg/dl) for 48 h. The collected cells were then fixed using cold 70% ethanol at −20 °C over 24 h, washed with PBS and incubated with 50 μg/mL propidium iodide (Becton, Dickinson and Company, Franklin lake, NJ, USA) at room temperature for 15 min. After being supplemented with propidium iodide, cycle cell analysis was performed using EPICS XL (Beckman Coulter). Percentage of cells at each cell cycle phases was determined with Modfit LT Software.

Ke B., Zeng Y., Zhao Z., Han F., liu T., Wang J., Khalique A., Lu W.J., Chong J., Lan F, & He H. (2018). Uric acid: a potent molecular contributor to pluripotent stem cell cardiac differentiation via mesoderm specification. Cell Death and Differentiation, 26(5), 826-842.

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Tumor Dissociation and Single-Cell Isolation

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Primary and secondary tumors were weighed and digested for 20 min at 37 °C in 5 ml PBS plus MgCl₂/CaCl₂ (Gibco; Thermo Fisher Scientific) supplemented with 50 μg/mL DNase I (New England BioLabs) and 120 μg/mL Liberase solution (Roche Life Science). After the incubation, tumor pieces were mechanically ground through a 70-μm strainer (Falcon) and filtered through a 30-μm pre-separation filter (Miltenyi Biotech). Lymph nodes were squeezed through a 70-μm strainer. Red blood cell (RBC) lysis was performed using 1× RBC lysis buffer (eBioscience).

Onyshchenko K., Luo R., Rao X., Zhang X., Gaedicke S., Grosu A.L., Firat E, & Niedermann G. (2024). Hypofractionated radiotherapy combined with lenalidomide improves systemic antitumor activity in mouse solid tumor models. Theranostics, 14(6), 2573-2588.

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Mouse Spleen Isolation and Cell Counting

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C57BL/6 mice were sacrificed, and spleens were extracted for our experiment. Spleens were weighed and minced into small pieces (< 0.1 cm²) with a scalpel blade. Spleen pieces were resuspended in PBS and vortexed to further mince down. The excised spleen was transferred to a 30 μm pre-separation filter (Miltenyi Biotec, Bergisch Gladbach, Germany) for straining. The specimen was washed several times with PBS and ground up further to improve yield. Following a final centrifuge and PBS wash step, the specimen was resuspended in PBS and counted on the Countess II FL (Thermo Fisher, Waltham, MA).

Bou-Samra P., Chang A., Arambepola S., Guo E., Azari F., Kennedy G., Segil A, & Singhal S. (2023). Preclinical Lymph Node Model for Intraoperative Molecular Imaging of Cancer. Research Square.

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Flow Cytometric Analysis of Differentiated EBs

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Differentiated EBs at different time points were trypsinized for 2 min, dissociated with gentle agitation and passed through a 30 μm pre‐separation filter (Miltenyi Biotec Inc., Auburn, CA) to produce a single cell suspension. Cells (1 × 10⁶) were then stained with primary antibodies as follows: EphB4 at 1:100 dilution (R&D Systems, Minneapolis, MN), ephrinB2 at 1:100 dilution (R&D Systems), and rat IgG2a (control antibodies) at 1:100 dilution. PE‐ or APC‐conjugated secondary antibodies were used for flow cytometric analysis. Dead cells were gated out by a DNA dye (7‐Amino‐Actinomycin D, 7AAD staining). Isotype‐matched control antibodies were used to determine the background staining. All antibodies were purchased from BD Biosciences (Franklin Lakes, NJ). The cells were analyzed on FACSCalibur (BD Biosciences) with CellQuest software. Data analysis was performed using CellQuest or FlowJo Software.
Cell sorting was performed using Fluorescence Activated Cell Sorting (FACS) on FACSAria II (BD Biosciences).

Chen K., Bai H., Liu Y., Hoyle D.L., Shen W., Wu L, & Wang Z.Z. (2015). EphB4 Forward‐Signaling Regulates Cardiac Progenitor Development in Mouse ES Cells. Journal of Cellular Biochemistry, 116(3), 467-475.

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