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HALO®-96 Research
HALO®-96 PREP
HALO®-96 PCA^EQ

HALO®-96 Research

Assays for Human Lympho-Hematopoietic Stem and Progenitor Cell Research

Buy HALO®-96 Research assay kits

Available as assay kits and
contract research services.

HALO®-96 Research Applications

For all human and animal lympho-hematopoietic research applications.
Detect and measure proliferation of human and animal stem and progenitor cells.
Measure and compare proliferation status and/or potential between tissues and cell populations.
Experimental transplantation models.
Effects of growth factors and/or cytokines.
In vivo to in vitro assays.
Gene targeting assays.

Advantages of using HALO®-96 Research

HALO®-96 Research is the only assay system available to detect and measure lympho-hematopoietic proliferation of human and animal cells. (The colony-forming cell, CFC, does not measure proliferation; it is a differentiation assay because colonies are identified by the differentiation and maturation of the cells that form the colonies. Please see CAMEO™-4 and CAMEO™-96). See HALO® and CFC Information Flyer.
Distinguish between multiple primitive and mature stem cell populations and lineage-specific populations.
Reliably compare your results from one experiment to another and over time by calibrating and standardizing the assay (reagents provided).
Provides all the flexibility you need for your research, including adding your own growth factors and/or cytokines.
HALO® utilizes Suspension Expansion Culture (SEC) and bioluminomics™ Technology that can be easily multiplexed with other assay readouts to obtain the most information from the same sample.

Species

HALO®-96 Research is available for the following species:

Human
Non-human primate (Macaca fascicularis (Cynomologus) and Macaca mulatta (Rhesus))
Horse
Pig
Sheep
Dog
Rat
Mouse

Individual Human and Animal Cell Populations Detected using HALO®-96 Research (see Lympho-Hematopoietic System to view cell populations and growth factor/cytokine cocktails)

The following proliferation status or potential can be detected for individual populations using HALO®-96 SPCA:

Stem Cells

• HPP-SP 1: IL-3, IL-6, SCF, TPO, Flt3-L

• HPP-SP 2: EPO, GM-CSF, IL-2, IL-3, Il-6, IL-7, SCF, TPO, Flt3-L

• CFC-GEMM 1: EPO, GM-CSF, IL-3, IL-6, SCF, TPO, Flt3-L

• CFC-GEMM 2: EPO, GM-CSF, IL-3, IL-6, SCF, TPO

• CFC-GEM 1: EPO, GM-CSF, IL-3, IL-6, SCF

• CFC-GEM 2: EPO, GM-CSF, IL-3, SCF

• CFC-GEM 3: EPO, GM-CSF, G-CSF, IL-3, SCF

Progenitor Cells

• BFU-E 1: EPO, IL-3, SCF

• BFU-E 2: EPO

• GM-CFC 1: GM-CSF, IL-3, SCF

• GM-CFC 2: GM-CSF, G-CSF, IL-3, SCF

• GM-CFC 3: GM-CSF

• Mk-CFC 1: TPO, IL-3, SCF

• Mk-CFC 2: TPO

Lymphopoietic cells

• T-CFC: IL-2

• B-CFC: IL-7

No growth factors: Add growth factors/cytokines of choice.

Multiple Human and Animal Cell Populations Detected Simultaneously Using HALO®-96 Research

4-Population Assay

CFC-GEMM 1
BFU-E 1
GM-CFC 1
Mk-CFC 1

7-Population Assay

HPP-SP 2
CFC-GEMM 1
BFU-E 1
GM-CFC 1
Mk-CFC 1
T-CFC
B-CFC
+ Background control

Click image for larger view

Example of a 7-Population Comparison Between Human Tissues

The graph shows the response of 7 lympho-hematopoietic cell populations derived from mobilized peripheral blood CD34⁺ cells, bone marrow and umbilical cord blood mononuclear cells. Please note that whereas mobilized peripheral blood CD34⁺ cells are shown at 500 CD34⁺/well, bone marrow and umbilical cord blood are shown at 5,000 mononuclear cells/well.

Click image for larger view

Comparison of 7-Population Response between Species

The graph shows the response of 7 lympho-hematopoietic populations, HPP-SP 2, CFC-GEMM 1, BFU-E 1, GM-CFC 1, Mk-CFC 1, T-CFC, B-CFC and background, from sheep, pig, dog, rat and mouse bone marrow.

Use

For Research Use Only. Not for clinical diagnostic use

Major Equipment Required

Plate luminometer or multiparameter plate reader with "glow" luminescence measuring capability.

Kit Contents

HALO®-96 Research Master Mix
ATP standard
ATP high and low controls (not included with research kits)
ATP Enumeration Reagent
Sterile, 96-well plates
Non-sterile, 96-well plates
Sterile, adhesive foil covers
Instruction manual

HALO® Kit Manual

To download the HALO®-96 Research kit manual, please go to the Downloads Page.

Related Assays

HALO®-96 PREP

Assays to Detect Proliferation, Self-Renewal and Expansion Potential of Primitive Stem Cells.

Buy HALO®-96 PREP assay kits

Available as assay kits and
contract research services.

HALO®-96 PREP Applications

Designed specifically for the primitive lympho-hematopoietic cell population, HPP-SP (High Proliferative Potential - Stem and Progenitor Cell).
Uses secondary re-plating technology to substantiate the presence of primitive stem cells.
Faster and more easier LTC-IC assay alternative.
Experimental transplantation models.
In vivo to in vitro assays
Gene targeting assays.

Advantages of using HALO®-96 PREP for Your Research

The only commercially available assay that can detect and measure proliferation, self-renewal and expansion potential of primitive lympho-hematopoietic stem cells (HPP-SP).
Distinguish between quiescent and proliferating primitive stem cells.
Used to "prime" primitive stem cells out of quiescence and into cell cycle.
A faster, easier and lower-priced alternative to LTC-IC assays.
HALO® utilizes Suspension Expansion Culture and Bioluminomics™ Technology for the most accurate, sensitive and reliable assays available.
Reliably compare results from one experiment to another over time by calibrating and standardizing the assay.
Multiplex HALO®-96 PREP with flow cytometry or molecular expression analysis for even more information from the same sample.

Species

HALO®-96 PREP is available for the following species:

Human
Non-human primate
Horse
Pig
Sheep
Dog
Rat
Mouse

Cell Populations Detected

HPP-SP 1: Quiescent, primitive, lympho-hematopoietic stem cell population.
HPP-SP 2: "Fully stimulated" primitive lympho-hematopoietic stem cell population.

Use

For Research Use Only.

Major Equipment Required

Plate luminometer or multiparameter plate reader with "glow" luminescence measuring capability.

Kit Contents

HALO®-96 PREP Master Mixes
ATP standard
ATP enumeration reagent
Sterile, 96-well plates
Non-sterile, 96-well plates
Sterile, adhesive foil covers
Instruction manual

Related Assays

HALO®-96 PCA^EQ

Hematopoietic Progenitor Colony-Forming Unit (CFU) Equivalent Assays for Stem Cell Processing Laboratories

Buy HALO®-96 PCA^EQ assay kits

Available as assay kits and
contract research service.

HALO®-96 PCA^EQ Applications

An equivalent and alternative to, or replacement for, the "classic" colony-forming unit (CFU) assay for stem cell processing laboratories.
FDA, FACT and AABB alternative assays. (NMDP uses FACT and AABB standards).

NOTE: Although the community norm is to use the colony-forming unit (CFU) assay, such as CAMEO™-4, an equivalent and alternative assay, such as HALO®-96 PCA^EQ is acceptable by the FDA, since it satisfies "the requirements of the applicable statutes and regulations". In addition, AABB Standards for Cellular Therapy Services also allows "equivalent functional assays for any product", and the National Marrow Donor Program (NMDP) allows testing "consistent with AABB Standards".

Advantages of using HALO®-96 PCA^EQ in the Stem Cell Processing Laboratory

Transforms the "classic" and subjective CFC assay into an instrument-based, fully standardized and validated assay from which meaningful results regarding the proliferation ability of the cells can be obtained.
Incorporates the same growth factor/cytokine cocktails used for MethoCult® reagents for direct assay-to-assay comparison and correlation.
Faster, easier to use, less expensive and more accurate than any traditional CFU assay, even using automated colony counting.
HALO®-96 PCA^EQ utilizes Suspension Expansion Culture (SEC) and Bioluminomics™ Technology, the most advanced and cost-effective hematopoietic in vitro cellular functionality assay system available.

Tissues

HALO®-96 PCA^EQ can be used for the following tissues:

Mobilized peripheral blood
Umbilical cord blood
Bone marrow
Purified cell populations for any of the above tissues.

HALO®-96 PCA^EQ is used as an Alternative to or Replacement for CAMEO™-4 (using the same reagent) and MethoCult® (using the same growth factor cocktails)

The table below shows the HALO®-96 PCA^EQ assays that are equivalent to CAMEO™-4 assays and MethoCult® reagents used in the stem cell processing laboratory. The growth factor/cytokine cocktail is the same for all three.

Please note that for cord blood banks applying for a BLA, the FDA requires validation of all assays. Unlike HALO®-96 PCA^EQ, neither CAMEO™-4 nor MethoCult® reagents for the CFU assays have or include standards and controls and can therefore not be validated according to FDA regulations.

**CAMEO™-4 and HALO®-96 PCA^EQ Assays that are Equivalent to MethoCult® Reagents**
CAMEO™-4	HALO®-96 PCA^EQ	MethoCult®	Growth Factor/Cytokine Cocktail
KC-GEM2-50H	K2-PCA1-1,2,4*	H4434	EPO, GM-CSF, IL-3, SCF
KC-GEM3-50H	K2-PCA2-1,2,4*	H4034	EPO, GM-CSF, G-CSF, IL-3, SCF
K2-GM1-50H	K2-PCA3-1,2,4*	H4534	GM-CSF, IL-3, SCF
KC-GM2-50H	K2-PCA4-1,2,4*	H4035	GM-CSF, G-CSF, IL-3, SCF

*denotes that assay kits are available with 1, 2 or 4, 96-well plates

Click image for larger view

Replacing the CFU/CFC Assay with HALO®-96 PCA^EQ

The diagram to the left shows the results from HALO®-96 PCA^EQ and CAMEO™-4, performed in parallel for human bone marrow and umbilical cord blood mononuclear cells (MNC). Although proliferation can be measured on days 5, 6 or 7, the results shown are from day 7 for HALO®-96 PCA^EQ and colony counts from days 7 and 10 for CAMEO™-4. Results from day 7 for both assays can be directly compared with each other. Notice that equivalent results are obtained for both assays. Also notice that colony counts do not change significantly from day 7 to day 10. This is to be expected since once all of the colony-forming cells present in the culture start to proliferate, no new colonies will be formed. The existing colonies only grow in size due to proliferation of the cells. However, this information is lost to the user when working with a CFU/CFC assay. (Unless proper controls are included, automated colony counters and image analysis cannot provide quantitative data on cell proliferation in colonies). In contrast, cell proliferation increases exponentially over the first 10 days of culture and can be quantified using HALO®-96 PCA^EQ (See HALO®-96 SPC-QC to see how proliferation increases). The results clearly indicate that the "classic" CFU/CFC assay can be replaced by HALO®-96 PCA^EQ or any other HALO® assay, all of which provide quantitative, standardized and reliable results that can be compared over time. The equivalent HALO®-96 PCA^EQ assays for MethoCult® reagents are shown.