Introduction to Intracellular Staining Method for Flow Cytometry

Flow cytometry and immunostaining technology for intracellular protein biomarkers have opened up various new options for the research and clinical diagnosis of cell phenotypes and functional characteristics. In fact, the phenotypic identification of certain cell types requires the simultaneous labeling of both cell surface and intracellular markers. Furthermore, the cytoplasmic localization of typical membrane molecules has been proven to be the most reliable lineage marker—for instance, cytoplasmic CD3 and CD22 in undifferentiated leukemia. Therefore, intracellular immunostaining is essential in many cell biology and functional studies. To this end, Nordic-MUbio offers a range of advanced FIX&PERM® cell fixation/permeabilization reagent products, making the analysis of intracellular antigens as easy as that of cell surface antigens.

Introduction to FIX&PERM® Cell Fixation/Permeabilization Kit

The FIX&PERM® Cell Fixation/Permeabilization Kit contains two reagents: Fixation Reagent (Reagent A) and Permeabilization Reagent (Reagent B). First, Reagent A is used to fix cells in suspension; then, Reagent B is applied to permeabilize the cell membrane. This process enables antibodies to access intracellular structures while preserving the morphological scatter characteristics of the cells. Its specific formulation reduces background interference and allows for the simultaneous addition of the permeabilization reagent and fluorochrome-labeled antibodies, facilitating the staining of membrane proteins and intracellular components (such as cytoplasmic enzymes, nuclear proteins, oncoproteins, cytokines, and immunoglobulins). The FIX&PERM® Cell Fixation/Permeabilization Kit is compatible with all commercially available flow cytometers and can be used to analyze normal and malignant leukocyte populations from various human biological samples (e.g., blood, bone marrow).

Product List of FIX&PERM® Cell Fixation/Permeabilization Reagents

All products in the FIX&PERM® series comply with the CE certification standards.

Advantages of FIX&PERM® Cell Fixation/Permeabilization Reagents

ü Gently fixes cells while preserving their scatter characteristics for flow cytometry.

ü Enables simultaneous characterization of both intracellular and cell surface markers.

ü The entire process takes less than one hour; cells can be analyzed immediately or stored for 24 hours.

ü Implements rigorous quality control (QC) procedures: Each batch is tested using quality control blood samples, with scatter characteristics of leukocyte populations analyzed and compared to ensure consistency and reliability of results between batches.

ü Contains no volatile organic solvents, allowing transportation and storage at room temperature.

ü Compatible with a range of monoclonal antibodies targeting both cell membrane and intracellular antigens.

Application of FIX&PERM® Cell Fixation/Permeabilization Kit in Hematologic Malignancy Research

Hematologic malignancies, such as leukemia and lymphoma, are cancers originating from cells in blood-forming tissues like the bone marrow. Ki-67, an intracellular nuclear marker for cell proliferation, exhibits expression levels in patients with these hematologic cancers that hold certain clinical indicative value. In histopathology, Ki-67 is commonly detected using immunohistochemistry, while its application in flow cytometry is relatively limited—resulting in restricted detectable parameters among bone marrow cell populations from different sources.

Nies et al. utilized flow cytometry to analyze Ki-67 expression in cancer patients. By investigating Ki-67 expression during bone marrow hematopoiesis, monocytopoiesis, and the maturation stages of hematopoietic cells, they established reference values for different hematopoietic cell lineages in healthy individuals. The FIX&PERM® Cell Fixation/Permeabilization Kit was employed in combination with Ki-67 immunostaining and flow cytometry software to measure proliferative changes in cells across various maturation pathways. Consequently, reference values for this cell cycle parameter could be determined across different blood cell lineages and maturation stages.

As a supplier with 18 years of professional experience serving China’s life sciences sector, XMJ has earned the support and trust of numerous customers through its professional and high-quality services. Currently, it has become an authorized supplier for the official procurement platforms of R&D and production bases (in China) of multiple world-renowned multinational enterprises, pharmaceutical and diagnostic companies, as well as major universities and research institutions. Among its offerings, the FIX&PERM® Cell Fixation/Permeabilization Kit (distributed by XMJ under the Nordic-MUbio brand) is widely used in research institutions such as hospitals and universities. We have compiled a list of some antibody products compatible with the FIX&PERM® Cell Fixation/Permeabilization Kit. For more information about the FIX&PERM® Cell Fixation/Permeabilization Kit, please call XMJ’s national customer service hotline at 400-050-4006 or visit our official website.

List of Selected Mouse Anti-Human Monoclonal Antibodies Compatible with FIX&PERM® Cell Fixation/Permeabilization Kit

Nordic-MUbio is a well-known manufacturer of immunological reagents based in the Netherlands. On February 14, 2013, Nordic acquired MUbio and officially renamed itself Nordic-MUbio. In December of the same year, it acquired An Der Grub Bio Research GmbH (ADG), thereby adding the FIX&PERM® cell fixation/permeabilization reagent product line.

Nordic-MUbio adheres strictly to the ISO 9001 standard in producing high-quality antibodies, mainly including those targeting cell adhesion proteins, nuclear proteins, cytoskeletal proteins, and more. Over 80% of its antibodies have undergone frozen tissue immunohistochemistry testing, nearly 70% of its products have been validated by flow cytometry, and 50% of its products have passed Western Blot (WB) validation.

Meanwhile, Nordic-MUbio is also a pioneer in zebrafish antibody development, owning more than 50 types of antibodies applicable for zebrafish fluorescence detection. For more information about its immunological products, please call Ximeijie's national customer service hotline at 400-050-4006 or visit the official website.

Reference：

1. Nies, K.P.H. et al. Determination of the proliferative fractions in differentiating hematopoietic cell lineages of normal bone marrow. Cytometry A. 2018 93(11):1097-1105. PMID: 30176186.

2. Integration of the Ki-67 proliferation index into the Ogata score improves its diagnostic sensitivity for low-grade myelodysplastic syndromes. Leuk Res. 2022 Feb;113:106789. doi: 10.1016/j.leukres.2022.106789.

3. Optimized gating strategy and supporting flow cytometry data for the determination of the Ki-67 proliferation index in the diagnosis of myelodysplastic syndrome. Data Brief. 2022 Feb 22;41:107976. doi: 10.1016/j.dib.2022.107976.

4. Determination of the proliferative fractions in differentiating hematopoietic cell lineages of normal bone marrow. Cytometry A. (2018) 93, 1097-1105 . doi: 10.1002/cyto.a.23564