Mitochondria isolation kit for cultured cells

Manufactured by Abcam

Sourced in United Kingdom, United States

The Mitochondria Isolation Kit for Cultured Cells is a laboratory product designed to isolate mitochondria from cultured cells. It provides a standardized protocol and reagents to effectively separate mitochondria from other cellular components.

Automatically generated - may contain errors

Lab products found in correlation

Ab102536, by Abcam (3 mentions) Ab201111, by Abcam (3 mentions) Pi88700, by Thermo Fisher Scientific (3 mentions) Mitochondrial complex 3 activity assay kit, by Abcam (2 mentions) Complex 1 enzyme activity assay kit, by Abcam (2 mentions) Complete protease inhibitor cocktail, by Merck Group (2 mentions) Complex 1 enzyme activity microplate assay kit, by Abcam (2 mentions) Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions) Click go protein reaction buffer kit, by Vector Laboratories (1 mentions) Succinate colorimetric assay kit, by Merck Group (1 mentions) α ketoglutarate assay kit, by Merck Group (1 mentions) Halttm protease inhibitor cocktail, by Thermo Fisher Scientific (1 mentions) Complex 4 enzyme activity assay kit, by Abcam (1 mentions) Complex 2 enzyme activity assay kit, by Abcam (1 mentions) Nuclear and cytoplasmic protein extraction kit, by Beyotime (1 mentions) Glutaminase, by Abcam (1 mentions) Flotillin 2, by BD (1 mentions) Glial fibrillary acidic protein (gfap), by Cell Signaling Technology (1 mentions) Atp5a, by Abcam (1 mentions) Pvdf membrane, by Merck Group (1 mentions)

29 protocols using mitochondria isolation kit for cultured cells

Mitochondrial Isolation from Cultured Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cellular mitochondria were fractionated using Mitochondria Isolation Kit for Cultured Cells (Abcam, Cat# ab110170) by following instructions from the manufacturer.

Malik N., Kim Y.I., Yan H., Tseng Y.C., du Bois W., Ayaz G., Tran A.D., Vera-Ramirez L., Yang H., Michalowski A.M., Kruhlak M., Lee M., Hunter K.W, & Huang J. (2023). Dysregulation of mitochondrial translation caused by CBFB deficiency cooperates with mutant PIK3CA and is a vulnerability in breast cancer. Cancer research, 83(8), 1280-1298.

+ Open protocol

+ Expand

Mitochondrial Protein Synthesis Assay

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

The assay was performed as described previously (6 (link)). Briefly, 2 × 10⁷ WT and CBFB_KO MCF10A cells were plated in complete DMEM/F12 media. The next day, cells were transferred to a methionine-free complete DMEM/F12 media with 5% dialyzed horse serum. The cytosolic translation was blocked by treating cells with cycloheximide for 20 minutes. For the negative control, cells were treated with both cycloheximide (50 μg/ml) and chloramphenicol (150 μg/ml) for 50 minutes to block mitochondria protein translation. Subsequently, cells were labeled with 500 μM L-Homopropargylglycine for 4 hours. After incubation, were extracted using the Mitochondria Isolation Kit for Cultured Cells (Abcam, Cat# ab110170) as per the manufacturer’s instructions. Next, 70 μg of mitochondria protein lysate was click-labeled with 40 μM AZDye 594 Azide using Click-&-Go^™ Protein Reaction Buffer Kit (Click Chemistry Tools, Cat#1262) as per the manufacturer’s instructions. Subsequently, lysates were run on a NuPAGE gel, and the gel was scanned at FLA-6500 Starion (Fujifilm) to visualize fluorescent proteins.

+ Open protocol

+ Expand

Mitochondrial Respiratory Complex Analysis

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mitochondria were isolated from NRCMs using the Mitochondria Isolation Kit for Cultured Cells (ab110170, Abcam). The activities of complexes I and III in the mitochondrial ETC were assessed with the Complex I Enzyme Activity Assay Kit (ab109721, Abcam) and the Mitochondrial Complex III Activity Assay Kit (K520-100, Biovision) according to the manufacturer's instructions, respectively [44 (link)].
The contents of α-KG and Suc were measured using the α-Ketoglutarate Assay Kit (MAK054, Sigma) and the Succinate Colorimetric Assay Kit (MAK184, Sigma) in accordance with the manufacturer's instructions, respectively [44 (link)].

He H., Wang L., Qiao Y., Yang B., Yin D, & He M. (2021). Epigallocatechin-3-gallate pretreatment alleviates doxorubicin-induced ferroptosis and cardiotoxicity by upregulating AMPKα2 and activating adaptive autophagy. Redox Biology, 48, 102185.

+ Open protocol

+ Expand

Mitochondria Isolation from Cultured Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mitochondria were isolated from whole cells using the Mitochondria Isolation Kit for Cultured Cells (Abcam, United Kingdom). Briefly, cells were harvested and centrifuged at 1000 RCF for 5 min. Pellets were frozen at −80°C overnight to weaken membranes. Cells were defrosted, resuspended to 5 mg/mL in Kit Reagent A, and kept on ice for 10 min. Cells were then homogenized with 30 strokes of a Dounce homogenizer with pestle B, and then centrifuged at 1000 RCF for 10 min at 4°C. Supernatant was set aside, and the pellet resuspended in Kit Reagent B, homogenized with 30 strokes of a Dounce homogenizer with pestle B, and then centrifuged at 1000 RCF for 10 min. The supernatants were combined and centrifuged at 12,000 RCF for 15 min. The supernatant was discarded, and the pellet resuspended in 500 µL Kit Reagent C. Isolated mitochondria were quantified using a Bradford Assay, and their presence confirmed through staining with MitoTracker Deep Red with visual inspection under a fluorescence microscope (See Supplementary Figure S1).

Mould R.R., Kalampouka I., Thomas E.L., Guy G.W., Nunn A.V, & Bell J.D. (2023). Non-chemical signalling between mitochondria. Frontiers in Physiology, 14, 1268075.

+ Open protocol

+ Expand

Mitochondria Isolation from Cultured Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

The mitochondrial fractions were isolated using the Mitochondria Isolation Kit for Cultured Cells (Abcam) with some modifications. Cells were lifted with a cell lifter and centrifuged at 1000× g for 10 min at 4 °C. The cell pellets were resuspended to 5 mg/mL with reagent A, incubated on ice for 10 min, then sonicated and spun at 1000× g for 10 min. The supernatants were saved and the cell pellets were resuspended to the same concentration with reagent B, sonicated, and spun for 10 min at 4 °C. Finally, the two supernatants were thoroughly mixed and spun at 12,000× g for 15 min. The deposit (mitochondrial fraction) was resuspended with reagent C and kept at −80 °C until analysis.

Jassey A., Liu C.H., Changou C.A., Richardson C.D., Hsu H.Y, & Lin L.T. (2019). Hepatitis C Virus Non-Structural Protein 5A (NS5A) Disrupts Mitochondrial Dynamics and Induces Mitophagy. Cells, 8(4), 290.

+ Open protocol

+ Expand

Mitochondrial Isolation from NSCLC Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

The mitochondrial fraction was isolated using Mitochondria Isolation Kit for Cultured Cells (ab110171, Abcam, Boston, MA, USA) according to the manufacturer’s instructions. Briefly, NSCLC cells were washed with a pre-cooled PBS buffer three times and pelleted by centrifugation at 200× g for 5 min. The cells were frozen and thawed for three rounds to weaken cell membranes. Then the cells were suspended in Reagent A for incubation on ice for 10 min. Then the cellular suspension was homogenized in a pre-cooled Dounce homogenizer. The homogenates were centrifuged at 1000× g for 10 min at 4 °C and saved as supernatant #1. The pellet was resuspended in Reagent B, and the process was repeated to aid in cell rupturing. The homogenates were centrifuged and saved as supernatant #2. Supernatants #1 and #2 were combined and centrifuged at 12,000× g for 10 min at 4 °C. The pellet was suspended into Reagent C, supplemented with protease inhibitors (HaltTM Protease Inhibitor Cocktail, Thermo Fisher Scientific). The aliquots were frozen at −80 °C until use.

Xiao W., Ahluwalia P., Wang L., Howard J., Kolhe R., Rojiani A.M, & Rojiani M.V. (2022). TIMP-1 Dependent Modulation of Metabolic Profiles Impacts Chemoresistance in NSCLC. Cells, 11(19), 3036.

+ Open protocol

+ Expand

Mitochondria Isolation from Human Platelets

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Human platelets were obtained from Bioreclamation (Westbury, NY, USA) from 500 ml of whole blood divided equally in 10 vials. The mitochondria was isolated as previously described [18 (link)] using a Mitochondria Isolation Kit for Cultured Cells from Abcam and Miltenyi Biotec.

Singh N.S., Habicht K.L., Moaddel R, & Shimmo R. (2017). Development and characterization of mitochondrial membrane affinity chromatography columns derived from skeletal muscle and platelets for the study of mitochondrial transmembrane proteins. Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 1055-1056, 144-148.

+ Open protocol

+ Expand

Mitochondrial Fractionation and STAT3 Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Mitochondria and cytosolic fractions from ispinesib naïve and resistant murine lines were isolated (Mitochondria Isolation Kit for Cultured Cells, Abcam, cat# 110170). Protein concentration was measured with a BCA assay (and fractions were stored at −80°C until further use. Samples were subjected to Western blotting using anti-pS727-STAT3, anti-STAT3, anti-cytochrome oxidase type IV, and anti α-tubulin antibodies.

Johnston J.A., Dalton M.J., Gurney M.E, & Kopito R.R. (2000). Formation of high molecular weight complexes of mutant Cu,Zn-superoxide dismutase in a mouse model for familial amyotrophic lateral sclerosis. Proceedings of the National Academy of Sciences of the United States of America, 97(23), 12571-12576.

+ Open protocol

+ Expand

Mitochondrial Enzyme Activity Assays

Check if the same lab product or an alternative is used in the 5 most similar protocols

Herein, mitochondria in cell samples were isolated using Mitochondria Isolation Kit for Cultured Cells (ab110170, Abcam, USA). Then, the activity of Complex I to IV was evaluated using Complex I Enzyme Activity Assay Kit (ab109721, Abcam, USA), Complex II Enzyme Activity Assay Kit (ab109908, Abcam, USA), Mitochondrial Complex III Activity Assay Kit (K520-100, Biovision, USA), and Complex IV Enzyme Activity Assay Kit (ab109909, Abcam, USA) according to the manufacturer’s instructions respectively.

Feng Y., Xu J., Shi M., Liu R., Zhao L., Chen X., Li M., Zhao Y., Chen J., Du W, & Liu P. (2022). COX7A1 enhances the sensitivity of human NSCLC cells to cystine deprivation-induced ferroptosis via regulating mitochondrial metabolism. Cell Death & Disease, 13(11), 988.

+ Open protocol

+ Expand

Mitochondria and Cell Fractionation

Check if the same lab product or an alternative is used in the 5 most similar protocols

The mitochondria of HEK293T cells were isolated using the Mitochondria Isolation Kit for Cultured Cells (Abcam), and the cytoplasm and nucleus of HEK293T cells were fractionated using the Nuclear and Cytoplasmic Protein Extraction Kit (Beyotime).

Zheng X., Wang M., Liu S., Chen H., Li Y., Yuan F., Yang L., Qiu S., Wang H., Xie Z, & Xiang M. (2023). A lncRNA-encoded mitochondrial micropeptide exacerbates microglia-mediated neuroinflammation in retinal ischemia/reperfusion injury. Cell Death & Disease, 14(2), 126.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!