AffiEXTRACT® Single Nuclei Isolation Kit

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CAT# AFG-INV-25
Size: 20 Isolations

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Sample Types

Suitable for fresh/frozen tissues and cultured cells.

Processing Time

Complete nuclei isolation in approximately 30 minutes.

Storage Conditions

Long term at +4°C and up to week at RT.

Yield 

10^6 nuclei per 10 mg of soft tissue.

Sample Size Range

Works with up to 25 mg of tissue or up to 10^7 cells

Buffer Components

Includes proprietary lysis and wash solutions optimized for efficient nuclei release.

Centrifugation Requirements 

Operates at 500-800 X g, compatible with standard benchtop centrifuges.  

Compatibility  

Effective across a wide range of tissue types; specialized for high-purity, single nuclei isolation.


Quick Protocol Guide


Collect/Wash: Centrifuge cells; wash with Nuclei Preservation Buffer.

Lysis: Add lysis solution; grind; incubate on ice.

Filter: Pour lysate into filters; centrifuge.

Wash/Resuspend: Clean with wash solution; resuspend in stabilizing buffer.


The "AffiEXTRACT® Single Nuclei Isolation Kit" is designed to isolate individual nuclei from various types of cells and tissues, making it ideal for downstream applications such as single-nucleus RNA sequencing (snRNA-seq). Here's how the kit works:


Step-by-Step Process:


1. Sample Preparation:

A small sample of tissue or a collection of cells is prepared for processing. This kit is suitable for a wide range of tissues and cell types, allowing you to isolate nuclei even from challenging samples.

2. Tissue and Cell Dissociation:

The tissue or cell sample is gently broken down into smaller parts. This step is crucial for releasing the nuclei from the sample while keeping them intact. The dissociation process is carefully designed to preserve the integrity of the nuclei for high-quality analysis.

3. Nuclei Isolation:

The kit uses specialized reagents that help break open the cell membranes while leaving the nuclei intact. This ensures that the nuclei are separated cleanly from the rest of the cell material, ready for further study.

4. Nuclei Purification:

After isolation, the nuclei are purified to remove debris and other unwanted components from the sample. The kit ensures that the nuclei are clean and intact, making them suitable for high-quality downstream applications.

5. Quality Check:

The number and quality of the isolated nuclei are checked. Researchers can use a microscope or automated counting device to ensure the nuclei are intact and suitable for further use in sequencing or other applications.

6. Downstream Applications:

Once isolated, these nuclei are perfect for various applications, including:

  • Single-nucleus RNA sequencing (snRNA-seq): Ideal for tissues where isolating whole cells is difficult.
  • Genomic and proteomic studies: Researchers can perform detailed analyses of the isolated nuclei to study gene expression and protein content.


Why Use This Kit for Cells and Tissues?


  • Versatile: This kit works well with a wide range of cells and tissues, making it suitable for many types of research.
  • High-Quality Nuclei: The kit ensures that the isolated nuclei are intact and pure, which is critical for accurate downstream analysis.
  • Ideal for Difficult Samples: In cases where isolating whole cells is challenging, this kit allows researchers to focus on nuclei, providing a more feasible approach.

Applications in Research:


AffiEXTRACT® Single Nuclei Isolation Kit is an advanced tool designed to efficiently isolate and purify nuclei from tissues and cultured cells, overcoming common challenges like aggregation and doublets. Its versatile applications make it a valuable resource across various scientific research fields:

Single Nucleus Sequencing (RNA-seq and ATAC-seq)

The kit is essential for preparing high-quality nuclei for single nucleus RNA sequencing and chromatin accessibility assays (ATAC-seq), crucial for understanding gene expression and chromatin dynamics in heterogeneous tissues at a single-cell resolution.

Epigenetic Studies:

- ChIP-seq (Chromatin Immunoprecipitation Sequencing): The isolated nuclei can be used to study protein-DNA interactions, helping researchers explore epigenetic regulation and modifications such as histone modifications.

- DNA Methylation Analysis: This enables the examination of methylation patterns across different conditions, providing insights into gene regulation, imprinting, and disease mechanisms.

Nuclear Proteomics:

Mass Spectrometry: Researchers can utilize isolated nuclei to identify and quantify nuclear proteins, facilitating studies on nuclear functions, including transcription, DNA repair, and replication.

Drug Screening and Pharmacological Studies:

Nuclear Response to Drugs: The kit can be used to study the effects of various drugs on nuclear functions, including how compounds affect DNA repair, nuclear signaling pathways, or gene transcription, making it valuable in drug development and testing.

Nuclear Morphology and Structural Studies:

Electron Microscopy: The isolated nuclei are suitable for detailed visualization using electron microscopy, allowing for in-depth analysis of nuclear structure, chromatin organization, and overall nuclear architecture.

Transcriptomics & Gene Expression:

- qPCR and RT-qPCR: RNA extracted from the isolated nuclei can be used in quantitative PCR studies, enabling precise measurements of gene expression within the nucleus.

- In Situ Hybridization: Enables detection of specific nucleic acid sequences, facilitating the study of gene localization and expression directly within nuclear compartments.

Flow Cytometry (FACS) Analysis:

The kit prepares nuclei for use in flow cytometry, enabling sorting and analysis based on nuclear characteristics, which is vital for cell population studies, including tumor profiling and stem cell research.

Immunofluorescence Staining:

Researchers can stain the isolated nuclei with fluorescent markers, allowing visualization and study of nuclear proteins and structures, supporting experiments related to protein localization and nuclear compartmentalization.

Cell Cycle Analysis:

The nuclei isolated with this kit can be used to study the different phases of the cell cycle, providing insights into cell division, growth, and proliferation, which are crucial in cancer research.


In summary, the AffiEXTRACT® Single Nuclei Isolation Kit provides a reliable way to isolate high-quality nuclei from a variety of samples. This makes it an essential tool for researchers who need to perform detailed genetic or cellular analysis, particularly in challenging tissues.