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PKH Fluorescent Cell Linker Kits

Catalog
Number

Description
Click on Product Name for additional catalog information

Product Information Sheet Available

PKH67-GL

PKH67 Green Fluorescent Cell Linker Kit
For general cell membrane labeling

PKH2-GL

PKH2 Green Fluorescent Cell Linker Kit
For general cell membrane labeling

PKH2-PCL

PKH2 Green Fluorescent Cell Linker Kit
For phagocytic cell labeling

PKH26-GL

PKH26 Red Fluorescent Cell Linker Kit
For general cell membrane labeling

PKH26-PCL

PKH26 Red Fluorescent Cell Linker Kit
For phagocytic cell labeling

CCPS-1

The Cell Census Plus^TM System

GL-DIL

Diluent A Kit for PKH2-GL

PCL-DIL

Diluent B Kit for PKH2-PCL and PKH26-PCL

CGL-DIL

Diluent C Kit for PKH26-GL and PKH67

U.S. Patent Numbers: 4,783,401; 4,762,701; 4,859,584. Distributed for Phanos Technologies.

Catalog Information

PKH67 Green Fluorescent Cell Linker Kit, Catalog No. PKH67-GL

is for general cell membrane labeling. It has a longer aliphatic carbon tail than PKH1 and PKH2, two other green dyes previously described for in vitro and in vivo cell tracking.^1-4 Based on the longer tail length, in-house studies have consistently shown reduced cell-cell transfer for PKH67 as compared to PKH2.⁵

Slow loss of fluorescence has been observed in in vivo studies using PKH1 and PKH2.^1,2 PKH67 may exhibit similar properties since this behavior appears to be characteristic of green cell linker dyes, but not red cell linker dyes.⁵ Correlation of in vitro cell membrane retention with in vivo fluorescence half life in non-dividing cells predicts an in vivo fluorescence half life of 10-12 days for PKH67.⁵ Other green cell linker dyes with similar half lives have been used to monitor in vivo lymphocyte and macrophage trafficking over periods of 1-2 months,^1,2 suggesting that PKH67 will also be useful for in vivo tracking studies of moderate length.

Each kit contains 0.5 ml of PKH67 cell linker and 6 vials of Diluent C.

NEW PKH67 is now available in a sample size kit, containing 0.1 ml PKH67 Cell linker and 1 vial Diluent C. Catalog No. MINI-67

PKH2 Green Fluorescent Cell Linker Kit, Catalog No. PKH2-GL

is for general cell membrane labeling. It has been characterized in a number of model systems and has been found to be useful for in vitro cell labeling, in vitro proliferation studies and short term, in vivo cell tracking.^6-9 The half-life for elution of PKH2 from labeled rabbit red blood cells is 10-11 days. This slow elution may complicate interpretation of experiments assessing in vivo proliferation.

Each kit contains 0.5 ml of PKH2 cell linker and 6 vials of Diluent A.

PKH2 Green Fluorescent Cell Linker Kit, Catalog No. PKH2-PCL

is for phagocytic cell labeling. It is used to selectively label cells with phagocytic capabilities such as monocytes, macrophages, or neutrophils.^1,10,11 The labeling occurs through the formation of dye aggregates or particulates. The aggregate formation significantly inhibits the uptake of dye by non-phagocytic cells, such as lymphocytes, but facilitates dye uptake by phagocytic cells. Labeled cells appear patchy or spotted because the dye is localized in phagocytic compartments of the cells. The dye appears to be resistant to metabolic attack and has been found to remain with the cells for more than 21 days in vivo.

Labeling of phagocytic cells by this methodology may be conducted either in vitro or in vivo. Intraperitoneal or intravenous injections of the PKH2 labeling solution will successfully label phagocytic cells in vivo, while cells of interest which have been isolated may be stained using in vitro labeling methods.

Each kit contains 0.5 ml of PKH2 cell linker and 6 vials of Diluent B.

PKH26 Red Fluorescent Cell Linker Kit, Catalog No. PKH26-GL

is for general cell membrane labeling. It has been characterized in a number of model systems and has been found to be useful for in vitro cell labeling, in vitro proliferation studies and long term, in vivo cell tracking.^6,9The half-life for elution of PKH26 from labeled rabbit red blood cells is greater than 100 days. This enhanced stability is favorable for long term in vivo studies.

Each kit contains 0.5 ml vial of PKH26 cell linker and 6 vials of Diluent C.

NEW PKH26 is now available in a sample size kit, containing 0.1 ml PKH26 Cell linker and 1 vial Diluent C. Catalog No. MINI-26

PKH26 Red Fluorescent Cell Linker Kit, Catalog No. PKH26-PCL

is for phagocytic cell labeling. It is used to selectively label cells with phagocytic capabilities such as monocytes, macrophages or neutrophils.^1,10,11The labeling occurs through the formation of dye aggregates or particulates. The aggregate formation significantly inhibits the uptake of dye by non-phagocytic cells, such as lymphocytes, but facilitates dye uptake by phagocytic cells. Labeled cells appear patchy or spotted because the dye is localized in phagocytic compartments of the cells. The dye appears to be resistant to metabolic attack and has been found to remain with the cells for at least 21 days in vivo.

Labeling of phagocytic cells by this methodology may be conducted either in vitro or in vivo. Intraperitoneal or intravenous injections of the PKH26 labeling solution will successfully label phagocytic cells in vivo, while cells of interest which have been isolated may be stained using in vitro labeling methods.

Each kit contains 0.5 ml of PKH26 cell linker and 6 vials of Diluent B.

References

1. Melnicoff, M.J., et al., J. Leuk. Biol., 44, 367 (1988).

2. Teare, G., et al., Cell Immunol., 134, 157 (1990).

3. Slezak, S., and Horan, P., J. Immunol. Meth., 177, 205 (1989).

4. Unpublished data, Zynaxis, Inc. and Sigma Chemical Co.

5. Unpublished data, Zynaxis, Inc.

6. Slezak, S., and Horan, P., Blood, 74, (6), 2171 (1990).

7. Samlowski, W., et al., J. Immunol. Methods, 144, 101 (1991).

8. Hugo, P., et al., Nature, 360, 679 (1992).

9. Messina, L., et al., Proc. Nat. Acad. Sci. USA, 89, (24), 12018 (1992).

10. Melnicoff, M.J., et al., J. Leuk. Biol., 43, 387 (1988).

11. Melnicoff, M.J., et al., Cellular Immunol., 118, 178 (1989).

CCPS-1 The Cell Census Plus^TM System, Catalog No. CCPS-1

The Cell Census Plus System is a cell proliferation assay for flow cytometry. Using flow cytometry, Cell Census Plus simultaneously measures cell proliferation of phenotypically distinct subsets of cells within a heterogeneous population.

CELL CENSUS PLUS^TM:

• Provides the number of cell divisions (up to ten generations), relative abundance of each daughter generation, the fraction of non-proliferating cells and the proliferation index. PLUS
• Study distinct cell subsets within a total cell population via immunophenotyping (i.e., antibodies to cell surface markers).
• Measures the endpoint of multistep activation, instead of discrete, early events, like up regulation of CD69.
• Provides for standardized cytometer setup and normalized cell counts.
• Works with ModFit (sold separately), a computer program, to deconvolute histograms.
• Labels all cell types with a universal fluorescent membrane dye. Membrane labeling is fast, and sufficient for kinetic studies.
• Does not interfere with biological and proliferative activities of cells.
• Uses nonradioactive reagents.

Cell Census Plus contains a red fluorescent dye to label the cell's membrane, microbeads to standardize instrument setup and normalize cell counts and software, the cell proliferation model, to interpret the histogram data.

The Cell Census Plus System takes advantage of a patented fluorescent aliphatic reporter molecule that acts as a membrane dye.¹ Its rapid uptake and stable retention in the plasma membrane and equal distribution into the membranes of the daughter cells make this dye ideal for cell tracking^2-5 and measuring proliferation. Cell labeling does not interfere with cell proliferation. Labeled cells will have a characteristic fluorescence intensity at time zero. Cell Census Plus assumes fluorescent intensity will reduce by one half at each cell division and each daughter cell will retain half the dye present in the mother cell. Therefore, the intensity will be 1/2 in the first daughter generation, 1/4 in the second daughter generation, 1/8 in the third daughter generation, etc.

Cell Census Plus can utilize immuno-gating techniques to measure proliferation of a select subset of cells within the proliferating cell culture.

The Cell Census Plus process is simple. First, label the entire cell population before exposing it to the proliferation stimulus. The cells are now ready for culture. After proliferation, the cells may be labeled with specific antibodies so that the proliferating activities of specific subpopulations may be studied. Next, ModFit (Verity Software) is used to analyze the data. ModFit is a computer program that deconvolutes raw histogram data into peaks. Each peak represents a generation. The Cell Proliferation Model software provides the parameters for ModFit to perform the histogram deconvolution. It then calculates the proliferation index, a ratio of the total number of cells analyzed versus the calculated number of parental cells required to generate the observed cell number. ModFit and The Cell Proliferation Model program can deconvolute histograms for up to ten generations and give the following numerical values: Number of cell divisions, Percent of cells in each generation for up to 10 generations, Proliferation index based on these percentages, Non-proliferating fraction, and Reduced chi-square as a measure of the goodness of fit.

To add quantitative strength to the results, use the PKH26 reference microbeads to standardize setup of the flow cytometer. Use of these microbeads ensures identical parameters for all assays. PKH26 reference beads are also used to normalize cell counts.

Each Kit Contains:

• 1 vial of PKH26 Dye, a red fluorescent membrane dye excited by the 488 nm argon ion laser found in most flow cytometers. The emission can be detected by using the standard filter setup for phycoerythrin (PE). Each vial of PKH26 dye contains 0.5 ml at 1 x 10^-3 M.
• 6 x 10 ml Diluent C, a patented dilution buffer. Cells are resuspended in Diluent C and PKH26 dye is diluted in Diluent C just prior to general surface membrane labeling. Diluent C is supplied sterile.
• 100 ml PKH26 Reference Microbeads, polymer microbeads having fluorescence intensity and light scatter characteristics similar to PKH26-labeled human leukocytes. The reference microbeads are impregnated with PKH26 dye, then carefully formulated as a suspension to serve as a fluorescence intensity and count standard.
• 1 each of The Cell Proliferation Model Software, a computer program for interpreting raw histogram data. The Cell Proliferation Model program runs with ModFit, (Product No. Z35,732-4) a DOS program that operates in Windows, and can analyze list mode files from a variety of flow cytometers. ModFit is available from Sigma and Verity Software House, Inc.

References

1. Horan, P., and Slezak, S., Nature, 340, 167 (1989).

2. Slezak, S., and Horan, P., Blood, 74, 2171 (1990).

3. Samlowski, W., et al., J. Immunol. Meth.,144, 101 (1991).

4. Hugo, P., et al., Nature, 360, 679 (1992).

5. Messina, L., et al., Proc. Natl. Acad. Sci. USA, 89, 12018 (1992).