"Tagging" Cells is Critical Past Methods Have Disadvantages PKH Fluorescent Dyes Offer Advantages Characteristics of PKH Dyes Methods for General Cell Labeling References/Product Information Achieve stable, uniform, intense, and reproducible fluorescent labeling of live cells with PKH kits from Sigma. Stable labeling for up to 100 days Non-cytotoxic with no effect on biological or proliferative activity Intense and easy-to-use Versatile -- use with any cell type Minimal leaking or transfer from cell to cell Able to combine with fluorescent antibodies or markers of cell function Compatible with existing detection equipment and standard fluorescence filters Technology that works, with literature to prove it Sigma is the only provider of PKH Fluorescent Cell Linker Kits for in vivo or in vitro cell tracking.

PKH Linker Kits for Fluorescent Cell Labeling

"Tagging" Cells Is Critical Many key research findings in cell function and pathogenesis have relied on labeling live cells. To study individual cell behavior, growth and differentiation, and cell-cell interactions, it is critical to be able to permanently "tag" a population of cells without affecting their morphology or function. Studies of cellular phenomena like adhesion, conjugate formation, cytotoxicity, phagocytosis, growth promotion, survival time, apoptosis, and hybridoma fusion all rely on dependable, nontoxic, stable cell labeling. Past Methods Have Disadvantages Early methods for labeling cells employed fluorescent labels such as fluorescein, rhodamine, DiI-C18-(3), or Hoechst dyes; or radiolabels like 125I-idoxuridine, 111In-indium oxine and 51Cr-sodium chromate. These methods served well in their time, but each has limitations and drawbacks.1 Some are not stable enough, so they leak and transfer to other cells in a mixed population, confounding results. Others alter cell function or are cytotoxic, and thus cause experimental artifacts related directly to their use. Finally, these methods often label weakly or non-uniformly, making detection difficult and design of reliable labeling protocols impossible.

Fluorescent micrographs showing the migration of PKH26 dye-labeled tumor-infiltrating lymphocytes in the spleen three days following intravenous injection of labeled live cells. (Photograph courtesy of L. Palmer & T. Schmitt, Zynaxis Cell Science, Inc.)

PKH Fluorescent Dyes Offer Advantages Now there is a cell labeling method that avoids all the pitfalls of these traditional methods. There is a single "best way" to consistently and reliably label viable cells: PKH Fluorescent Cell Linker Kits from Sigma (see Table 1).

Table 1 Desirable characteristics of cell labels. Notice that PKH dyes are the only class that meet all criteria.

Named for their discoverer, Paul Karl Horan, these patented fluorescent dyes and cell labeling technology2 have been used successfully with animal, plant, and bacterial cells, and even non-cellular membrane-containing particles.3 Labeled cells can be studied in culture and in vivo. Rapidly dividing cells like hybridomas, as well as non-proliferative cells like red blood cells, can be labeled and tracked. Easy to use. When you buy from Sigma, you get the PKH dyes, the specially formulated diluents, and the tested labeling protocols all in kit form. Weíve already determined the optimum combinations of reagents and conditions for intense, fast and reproducible viable-cell labeling in many model systems. Our kits make PKH labeling easy. Versatile. The versatility of PKH cell-labeling technology is attributable to its innovative chemistry. Intensely fluorescent dye moieties are attached to long, lipophilic tails. During the short general membrane staining procedure -- only 1-5 min. -- the lipophilic tails diffuse into the cell membrane, leaving the fluorogenic moiety exposed near the outer surface of the cell (Figure 1). Table 2 summarizes some common applications by cell type. Stable labeling without experimental artifacts. The stable partitioning of these molecules into the membrane permits long-term monitoring while leaving the important functional surface proteins unaltered. In fact, all cellular functions, from proliferation to cell-surface antigen recognition, are largely unaffected by PKH dye tagging. In short, labeled cells behave just like unlabeled ones; theyíre just easier to track. Intense, reproducible membrane staining. PKH dye labeling protocols can usually be optimized to give over 75% viable labeled cells with mean fluorescence more than 1,000 times background (see Figure 2). Such high signal-to-noise ratios are especially important when tagging proliferating cell lines because each daughter cell gets only half its parentís label. With PKH dyes, up to ten generations can be followed before the intensity of the label falls near background. Label by phagocytosis. Phagocytic cells can be selectively labeled in the presence of non-phagocytic cells by using an alternate diluent. In this diluent, dyes do not dissolve, but instead form microaggregates; these microaggregates cannot become embedded in membrane, but can be ingested by phagocytosis. This method has been used to study lifetime and migrational patterns of macrophages and neutrophils in vivo.

Table 2 Some documented applications of PKH dye labeling by cell type. For references to specific applications, call Technical Service.

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