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Organelle Isolation

Organelle Isolation Analysis Kits

Enriching for organelles simplifies proteomic analysis and when partnered with protein pathway function allows key insight into development of healthy cell function as well as the disruption that leads to disease. The kits and reagents shown below enable researchers to enrich for functional mitochondria, chloroplasts and more, with companion kits to determine organelle integrity.

Features and Benefits:

Isolation of organelles from different organisms - mammalian tissues and cultured cells, plant leaves and yeast cells

Specially formulated extraction reagents for research scale applications - save time and minimize waste

Produces functional intact organelles - suitable for functional studies, metabolic assays and protein profiling

Compatible with products for structure confirmation - easily confirm intactness with companion test kits

 

Product Name Product # Enrich for Recommended Antibodies for immunodetection* Companion test kits*
CelLytic™ NuCLEAR™ Extraction Kit NXTRACT Nucleus (mammalian tissue or cultured cells) L1293 - Anti-Lamin A
N2662 - Anti-Nucleolin
AMAB90549 - Anti-FUS
 
CELLYTIC™ PN ISOLATION/EXTRACTION KIT (For plant leaves) CELLYTPN1 Nucleus (plant) H0164- Anti-Histone H3  
LYSOSOME  ISOLATION  KIT LYSISO1 Lysosomes (animal tissue or cultured cells) L1418 - Anti-LAMP1
L0668 - Anti- LAMP2
C0715 - Anti-Cathepsin D
CS0740 - Acid phosphatase Assay Kit
MITOCHONDRIA ISOLATION KIT MITOISO1 Mitochondria (animal tissue) M6319 - Anti-Mitofusin 2 (N-terminal)
S5069 - Anti-MnSOD (DD-17)
T6319 - Anti-TOM22
C9616 - Anti-Cytochrome C
CYTOCOX1 - Cytochrome c Oxidase Assay Kit
CS0720 - Citrate Synthase Assay Kit
CS0390 - Mitochondria Staining Kit
CS0760 - Isolated Mitochondria Staining Kit
CELL MITOCHONDRIA ISOLATION KIT MITOISO2 Mitochondria (cells)
YEAST MITOCHONDRIA ISOLATION KIT MITOISO3 Mitochondria (yeast)  
ENDOPLASMIC RETICULUM ISOLATION KIT ER0100 Endoplasmic Reticulum (ER) G9043 - Anti-GRP78/BIP (ET-21)
P7496 - Anti-Protein disulfide isomerase (PDI)
CY0100 - Cytochrome c Reductase (NADPH) Assay Kit
CHLOROPLAST ISOLATION KIT CPISO Chloroplasts (plant leaves)    
CAVEOLEA/RAFTS ISOLATION KIT CS0750 Caveolae and lipid rafts (tissue culture cells) SAB4200216 - Anti-Caveolin 1
C3237 - Anti-Caveolin 1
C4490 - Anti-Caveolin 1
F1180 - Anti-Flotillin 1
 
GOLGI ISOLATION KIT GL0010 Golgi membranes (mammalian soft tissues) T7576 - Anti-TGN46
G7295 - Anti-GM130 (C-terminal)
 
PEROXISOMES ISOLATION  KIT PEROX1 Peroxisomes (animal tissues and cells) P0497 - Anti-PMP70
SAB4200181 - Anti-PMP70
C0979 - Anti-Catalase
CAT100 - Catalase Assay Kit

*Recommended antibodies and companion kits (enzymatic and colorimetric assays) to ensure the quality of organelle preparations.

 

CelLytic™ NuCLEAR™ Extraction Kit (NXTRACT)

For mammalian tissue or cultured cells

This kit features specially formulated extraction reagents and optimized protocols for the preparation of nuclear protein extracts from mammalian tissue or cultured cells (adherent or in suspension) using a hypotonic buffer that allows cells to swell. The cells are then disrupted, the cytoplasmic fraction is removed, and the nuclear proteins are released from the nuclei by a high salt buffer. This protocol applies for the extraction of functional, crude nuclear proteins from various cell types. Nuclear proteins can be extracted either from a small number of cells (106 -107 cells, 20-100 ml of packed cell volume), or from a much larger number of cells (108 -1010 cells, ~1 ml of packed cell volume). The kit is sufficient for 100 extractions of 100 ml of packed cell volume or 10 extractions of 1 ml of packed cell volume. The protein extracts are suitable for the detection of DNA-protein interaction using a gel-shift assay, DNase I footprinting analysis, and similar techniques.

CelLytic™ NuCLEAR™ Extraction Kit Protocol

Features & Benefits

  • Provides a complete system for preparing nuclear and cytoplasmic protein extracts from mammalian tissue or cultured cells.
  • Specially formulated extraction reagents and a proven procedure suitable for small scale isolation - Save time and increase confidence in the quality of the nuclear extract preparation.
  • Protocol tested for HeLa, CHO, COS, PC-12, Jurkat, and Bovine Aorta Endothelial Cells (BAEC).
  • Nuclear proteins can be extracted from the following fresh or frozen tissues: mouse brain and lung, rat liver, and rabbit muscle.
  • Suitable for both proteomic and genomic applications.

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CelLytic™ PN Isolation/Extraction Kit (CELLYTPN1)

For plant leaves

CelLytic™-PN extraction kit is for the isolation of nuclei and extraction of functional nuclear proteins from plant leaves. Nuclei or nuclear proteins can be extracted from few grams to hundreds grams of fresh and frozen leaves. The nuclear protein extract is suitable for the detection of DNA-protein interactions using gel-shift assay, DNase-I footprinting analysis as well as western blot assay and similar techniques. The isolated nuclei can also be used as a source for chromatin, genomic DNA and histones.

CelLytic™ PN Isolation/Extraction Kit Protocol

Features & Benefits

  • For the rapid isolation of nuclei and extraction of functional nuclear proteins from plant leaves.
  • Nuclei or nuclear proteins can be extracted from a few grams to hundreds of grams of fresh and frozen leaves.
  • Provides a detailed protocol for nuclei isolation and protein extraction from four plant models: tobacco, tomato, spinach, and Arabidopsis.
  • Suitable for both proteomic and genomic applications.

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Mitochondria Isolation Kit (MITOISO2)

For cultured cells

This kit enables the fast and easy isolation of an enriched mitochondrial fraction from cells. Most of the isolated mitochondria will contain intact inner and outer membranes. The outer membrane integrity may be measured by observing cytochrome c oxidase activity (CYTOCOX1) and the integrity of the inner membrane can be detected by the measurement of the citrate synthase activity (CS0720). The inner membrane integrity can also be assessed by testing the electrochemical proton gradient (ΔΨ) of the inner mitochondrial membrane by measuring the uptake of the fluorescent dye JC-1 into the mitochondria (CS0760 or T4069). In addition, the kit may be used for isolating mitochondrial proteins for proteome studies.

Mitochondria Isolation Kit Protocol

Features & Benefits

  • Specially formulated extraction reagents and a proven procedure suitable for small scale isolation - Isolate an enriched, intact mitochodrial fraction.
  • Produces functionally active, intact mitochondria - Resulting mitochondria are suitable for functional and proteome studies.
  • Compatible with the Cytochrome c Oxidase Assay Kit - Allows easy determination of the integrity of the outer mitochondrial membrane.
  • Tested on NIH 3T3, HeLa, HEK 293T, A431, HepG2, CHO, and Jurkat cell lines.

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Yeast Mitochondria Isolation Kit (MITOISO3)

For Yeast cells

The kit contains all the reagents required for yeast cell wall lysis by lyticase (spheroplast formation) followed by cell membrane lysis/breakage and mitochondria isolation. This kit enables the fast and easy yeast cell wall lysis, and isolation of an enriched mitochondrial fraction from yeast cells. The reagents are sufficient for 40 procedures using 20 OD culture preparations.

Yeast Mitochondria Isolation Kit protocol

Features & Benefits

  • Provides speed, sensitivity, ease of use, and specificity for mitochondria islation
  • Provides two alternatives for cell membrane disruption; using a detergent or by homogenization.
  • Includes an extraction buffer for mitochondrial protein profiling to be used in proteome studies
  • Tested on Saccharomyces cerevisiaePichia pastoris, and Schizosaccharomyces pombe
     

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Caveolea/Rafts Isolation Kit (CS0750)

For tissue culture cells

The Caveolae/Rafts Isolation Kit provides an easy method for obtaining fractions enriched with caveolea/rafts proteins from tissue culture cells. The method for the isolation of caveolae and rafts is based on the insolubility of these structures in ice cold TRITON- X-100. The kit is sufficient for 20 purifications.

Caveolea/Rafts Isolation Kit Protocol

Features & Benefits

  • Specially formulated extraction reagents and a proven procedure suitable for small scale isolation.
  • Includes all reagents for cells lysis, density gradient, and the isolation of caveolea/rafts protein enriched fraction, as well as two markers for caveolea/rafts compartments, anti-caveolin-1 and cholera toxin B subunit-peroxidase (CTB-HRP).
  • Tested on A431, HeLa, NIH 3T3, HdFn, HFF, BAEC, PC-12, BHK, HEK 293T, Jurkat, HL-60, NSO, K562, and U937 cell lines.

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Golgi Isolation Kit (GL0010)

For mammalian soft tissues

The Golgi Isolation Kit provides a method for isolating Golgi membranes from mammalian soft tissues by discontinuous density gradient. The degree of Golgi enrichment can be determined by assaying the acitivty of UDP-galactosyl transferase or by immunodetection of Golgi specific marker proteins like B-COP or GM130 using appropriate antibodies (G6160 and G7295, respectively). Separation from other organelles can be measured using the appropriate marker detection kits (CS0780, CYTOCOX1, CY0100 and CAT100). The kit is  sufficient for 50 g (tissue).

Golgi Isolation Kit protocol

Features & Benefits

  • Specially formulated extraction reagents and a proven procedure suitable for small scale isolation.
  • Optimized using rat liver and tested on rat kidney, spleen, and heart.

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Lysosome Isolation Kit, (LYSISO1)

For the enrichment of lysosomes from tissues and packed cells for functional and profiling studies

Lysosomes are organelles ubiquitously distributed in most pro- and eukaryotic cells. The Lysosome Isolation Kit provides a method (see protocol) for isolating lysosomes from animal tissues and from cultured cells by differential centrifugation followed by density gradient centrifugation and/or calcium precipitation (Lysosome Isolation Kit Data). The presence of lysosomes can be measured by assaying the activity of the lysosomal marker Acid Phosphatase, with the Acid Phosphatase Assay Kit (CS0740). Separation from other organelles can be measured using the appropriate marker detection kits listed below. The kit is sufficient for the preparation of lysosomes from 25 g of tissue or 20 ml of packed cells.


Lysosome Isolation Kit Protocol


Features & Benefits

  • Generate functional organelles for metabolism and disease research
  • Includes protease inhibitor to manage degradation
  • Measure integrity with included Neutral Red dye
  • Choose from multiple options to obtain highly enriched fractions

 

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Mitochondrial Isolation Kit, (MITO-ISO1)

For the isolation of an enriched, functional mitochondrial fraction from animal tissue

This kit features specially formulated extraction reagents and optimized protocols for the preparation of mitochondrial fractions from either soft (liver or brain) or hard (skeletal or heart muscle) tissue. In addition, the integrity of the inner membrane of the resulting mitochondria is analyzed by measuring the uptake of the fluorescent carbocyanine dye (JC-1) which is supplied in the kit (Mitochondrial Isolation Kit Data). The integrity of the mitochondrial outer membrane may be assessed by measuring cytochrome c oxidase activity using the Cytochrome c Oxidase Assay Kit (CYTOC-OX1). The resulting intact mitochondrial preparation is suitable for apoptosis studies, 2D analysis for proteomics and many other applications. Each kit is sufficient for the extraction of mitochondria from 10 to 20 grams of animal tissue. Enough JC-1 staining solution is provided to perform 50 JC-1 assays of 2 mL each.


Mitochondrial Isolation Kit Protocol


Features & Benefits

  • Specially formulated extraction reagents & a proven procedure suitable for small scale isolation - Isolate an enriched, intact mitochodrial fraction in a microfuge tube.
  • Produces functionally active, intact mitochondria - Resulting mitochondria are suitable for in vitro assays for apoptosis, oxidative stress or other studies.
  • Includes stain & protocol for determining inner membrane integrity - Determine the integrity of the inner mitochondrial membrane without the need to purchase other reagents.
  • Compatible with the Cytochrome c Oxidase Assay Kit - Allows easy determination of the integrity of the outer mitochondrial membrane

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Cytochrome C, (CYTOC-OX1)

For the determination of cytochrome c oxidase activity in soluble & membrane bound mitochondrial samples.

Cytochrome c oxidase is located on the inner mitochondrial membrane dividing the mitochondrial matrix from the intermembrane space. The Cytochrome c Oxidase Kit uses an optimized colorimetric assay based on observation of the decrease in absorbance of ferrocytochrome c measured at 550 nm, which is caused by its oxidation to ferricytochrome c by cytochrome c oxidase. This kit is suitable for the detection of mitochondrial outer membrane integrity and for the detection of mitochondria in subcellular fractions. (Cytochrome C Data). The Cytochrome c Oxidase Kit provides sufficient reagent for 100 tests.


Features & Benefits

  • Simple, optimized protocol - Obtain reproducible results without the need for special training.
  • Useful for determination of cytochrome c activity from any mitochondrial source - The enzyme is present in all mitochondria regardless of species.
  • Useful for detecting the presence of mitochondria in subcellular fractions - Save time & increase confidence in the quality of organelle preparations.
  • May be used in conjunction with the Mitochondrial Isolation Kit (MITO-ISO1) - Standardized mitochondrial preparation & analysis ensure reproducible results.

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Endoplasmic Reticulum Isolation Kit, (ER0100)

For the isolation of intact endoplasmic reticulum from mammalian soft tissues & cultured cell lines by centrifugal separation

The endoplasmic reticulum (ER) is a network of interconnected tubules, vesicles, and sacs. They play a role in specialized cellular functions including protein synthesis, sequestering of calcium, production of steroids, storage and production of glycogen, and insertion of membrane proteins. The kit contains all the reagents required for ER preparations with various degrees of purity: crude ER (microsomes), Ca2+ precipitated rough ER (RER) enriched microsomes, and density gradient purified rough ER (RER) and smooth ER (SER). The ER isolation process can be monitored by measuring the activity of NADPH cytochrome c reductase Cytochrome c Reductase Assay Kit, (Product Code CY0100). The enzyme is an ER membrane protein that is commonly used as an ER marker. The isolated ER can be used for studying the cytochrome P450 system and xenobiotic metabolism, for studying lipid metabolism, and for the recovery of ER membrane and lumenal proteins. The kit was tested with rat liver, kidney, and brain; mouse liver; rabbit liver, kidney, spleen, heart, and brain; and with Jurkat and HeLa cell lines.

Endoplasmic Reticulum Isolation Kit Flow Chart


Features & Benefits

  • Specially formulated extraction reagents & a proven procedure suitable for small scale isolation - Isolate an enriched, intact endoplasmic reticulum fraction in a microfuge tube.
  • Produces functionally active, intact endoplasmic reticulum(ER) - Resulting ER are suitable for in vitro assays for profiling, lipd metabolism, P450 studies & membrane protein analysis
  • Includes stain & protocol for determining inner membrane integrity - Determine the integrity of the endoplasmic reticulum membrane to confirm functional performance.
  • Compatible with the Cytochrome c Reductase Assay Kit, (CY0100) - Allows easy determination of the integrity of the outer endoplasmic reticulum membrane.

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Chloroplast Isolation Kit, (CP-ISO)

For the isolation of intact chloroplasts from plant leaves

This kit enables isolation of chloroplasts through mechanical cell wall & cell membrane breakage, removal of cell debris & unbroken leaf tissue by filtration, collection of total cell chloroplast fraction by centrifugation, & separation of intact chloroplasts (Data) from broken chloroplasts using a Percoll® layer or gradient. (Chloroplast Isolation Kit Data) Sigma's Chloroplast Isolation Kit has been tested for use with spinach, pea, lettuce, cabbage, mangold and tobacco.

Chloroplast Isolation Kit Protocol


Features & Benefits

  • Specifically formulated buffer - Saves time using easy-to-follow, simplified protocol based on proven methods.
  • Isolates intact chloroplasts - Useful for studies of such processes as carbon assimilation, electron flow and phosphorylation, metabolic transport, or protein targeting.
  • Kits are useful for both proteomic or genomic applications - Provides optimized conditions for isolation of intact chloroplasts from leaves of plants commonly used for research.

 

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Cytochrome c Reductase (NADPH) Assay Kit, (CY0100)

For the determination of cytochrome c oxidase activity in tissue and cell samples

NADPH-cytochrome c reductase is widely used as an endoplasmic reticulum marker and biomarker for ecological pollution and dietary lipid uptake. This kit is a fast and convenient analysis of NADPH-cytochrome c reductase activity in cell or tissue extracts and in purified microsomes [endoplastic reticulum (ER)]. It is suitable for samples prepared from mammalian tissues such as liver, kidney, brain, spleen, and heart muscle from several species, as well as on lysates from cell lines such as HeLa, HepG2, and Jurkat. Additionally, it has been tested on yeast strains P. pastoris and S. cerevisiae samples.

Features & Benefits

  • Detection of endoplasmic reticulum (ER) during isolation and subcellular fractionation by density gradient separation - Validates degree of enrichment and can be multiplexed to determine purity of enriched fraction against other organelles.
  • Assaying NADPH-cytochrome c reductase levels in various sources after intoxication by drugs or challenge by xenobiotics - Determine and compare the results of environmental perturbations.

 

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Peroxisome Isolation Kit, (PEROX1)

For the enrichment of intact peroxisomes from tissues and cells

Isolated peroxisomes are used for studying lipid b-oxidation, amino acid metabolism and biosynthesis of ether-linked glycerolipids and bile acids.This kit provides all the necessary reagents and a detailed protocol for the isolation of highly purified peroxisomes from animal tissues and cells, by differential density gradient centrifugation using iodixanol [Optiprep] density gradient centrifugation. The kit has been used for preparation of peroxisomes from rat liver, rat kidney and rabbit liver as well as HEK293 and HepG2 cells.

Features & Benefits

  • Specially formulated extraction reagents for research scale applications - Save time and minimize waste.
  • Produces functional intact organelles - Resulting peroxisomes are suitable for metabolic assays and disease state analysis.
  • Compatible with products for structure confirmation - Easily confirm intactness with companion kit test for cytochrome C reductase.

 

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Catalase Assay Kit, (CAT100)

For the determination of catalase activity in tissues and cells

A simple and easy colorimetric assay for analysis of peroxisome enrichment and catalase activity in various tissues. Catalase is a ubiquitous antioxidant enzyme in mammalian and non-mammalian aerobic cells containing a cytochrome system. Catalase provides protection against oxidative damage to living cells and it's role in oxidative stress related diseases has been widely studied.

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