HTRF® Anti-species reagents

Unique set of reagents for developing homogeneous high-throughput assays.

HTRF Protein-Protein interaction (PPI) reagents include secondary murine, sheep, rabbit, and human antibodies, as well as immunoglobulin binding proteins. These reagents are useful in developing new assays for which directly labeled assay components are not available or which involve chimeric proteins. Most of our anti-immunoglobulin reagents are available conjugated to either Europium-cryptate or XL665, for maximum flexibility.

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Product Specifications

Anti-Ig HTRF reagents have been strictly selected based on their specificity. Polyclonal antibodies (PAb) to murine, rabbit, and human immunoglobulins (IgGs) are absorbed against most human and animal serum proteins to minimize inter-species cross-reactivity.

Motif Active Moiety 5,000 tests
Europium conjugate XL665 conjugate
Protein A 3.5 µg 50 µg
Motif Active Moiety 5,000 tests Europium conjugate Active Moiety 5,000 tests XL665 conjugate
Protein A 3.5 µg 50 µg
Species Antibody Species Specificity Active Moiety
5,000 tests
Europium
conjugates
XL665
conjugates
Mouse immunoglobulins PAb Rabbit Mouse Igs 7.5 µg 100 µg
Rabbit immunoglobulins PAb Goat Rabbit Igs 7.5 µg 100 µg
Human immunoglobulins PAb Goat Human Fc Igs 10 µg 100 µg
Species Antibody Species Specificity Active Moiety 5,000 tests Europium conjugates Active Moiety 5,000 tests XL665 conjugates
Mouse immunoglobulins PAb Rabbit Mouse Igs 7.5 µg 100 µg
Rabbit immunoglobulins PAb Goat Rabbit Igs 7.5 µg 100 µg
Human immunoglobulins PAb Goat Human Fc Igs 10 µg 100 µg

Build HTRF assays for your specific application

HTRF PPI reagents provide an added level of flexibility to assay development, and are therefore commonly used when a ready-to-use assay kit is not available.

Reagents are sold by the number of tests (384-well, 20 µl reactions). Antibodies conjugated to XL665 or d2 are supplied on the basis of 20ng of antibody per well. The amount of active moiety per vial is also provided (as well as the number of tracers per vial - see product description sheet). In practice, the amount of active moiety is generally preferred to that of total conjugate as a basis for calculating assay development.

Recommended quantities of Eu3+ cryptate and XL665 conjugates

Most assays can be run within the nanomolar range. However, as a tracer, Eu3+ cryptate conjugates must not be excessive in order to prevent reader saturation and an unacceptable level of background. In most cases, a Eu3+ cryptate concentration of 4 to 5nM is appropriate, and will typically generate 620nm fluorescent signals of approximately 20,000 to 80,000 cps depending on the HTRF compatible reader used. As an example, for an antibody conjugated to Eu3+ cryptate with a molar ratio of 5 cryptates/Ab, the recommended value would be close to 1nM of antibody per well. The XL665 conjugate must match its assay counterpart as closely as possible in order for the maximum number of biomolecules to be tagged with the XL665 acceptor. Thus, to detect a GST-tagged molecule at an assay concentration of 20nM, the concentration of anti-GST-XL665 should be equimolar or higher. The actual amount will depend on the assay configuration.

Custom labeling and assay development services are also available to help meet your research needs.

Product Performances

CD28 provides a major costimulatory signal for CD4-positive T cells. Ligation with its natural ligands CD80 (B7-1) and CD86 (B7-2) leads to signals during activation that are required for the production of interleukin-2. This process has been implicated in the regulation of T-cell anergy and programmed cell death. The HTRF anti-immunoglobulin reagents were used to develop an assay for antagonists of CD28 and CD86 binding. Both molecules were respectively tagged with human and rat IgG Fc fragments, and detected using the XL665-labled anti-human antibody. CD86 was expressed as a fusion protein with a rat Ig domain which is recognized by a biotinylated sheep anti-rat antibody, followed by binding of europium-labeled streptavidin. Results of the binding assay were published by Mellor et al.

Data for the CD28-CD86 binding assay is published in: Mellor et al. Development of a CD28/CD86 (B7-2) binding assay for high throughput screening by homogenous time-resolved fluorescence. J Biomol Screening. 1998; 3: 91-99.

Part#, inserts & MSDS

Ordering Info

DescriptionCat. noProduct insertMSDS
PAb Anti Human IgG-d2 - 5,000 tests61HFCDAA
PAb Anti Human IgG-d2 - 20,000 tests61HFCDAB
PAb Anti Human IgG-d2 - 1,000 tests61HFCDAF
PAb Anti Human IgG-Eu cryptate- 5,000 tests61HFCKLA
PAb Anti Human IgG-Eu cryptate - 20,000 tests61HFCKLB
PAb Anti Human IgG-Tb cryptate - 5,000 tests61HFCTAA
PAb Anti Human IgG-Tb cryptate - 20,000 tests61HFCTAB
PAb Anti Human IgG-Tb cryptate - 1,000 tests61HFCTAF
PAb Anti Human IgG-XL665 - 5,000 tests61HFCXLA
PAb Anti Human IgG-XL665 - 20,000 tests61HFCXLB
PAb Anti Human IgG-XL665 - 1,000 tests61HFCXLF
MAb Anti Human Kappa-d2 - 5,000 tests61KAPDAA
-
MAb Anti Human Kappa-d2 -20,000 tests61KAPDAB
-
MAb Anti Human Kappa-Eu cryptate - 5,000 tests61KAPKAA
-
MAb Anti Human Kappa-Eu cryptate - 20,000 tests61KAPKAB
-
PAb Anti Mouse IgG-d2 - 5,000 tests61PAMDAA
PAb Anti Mouse-IgG-d2 - 20,000 tests61PAMDAB
PAb Anti Mouse IgG-d2 - 1,000 tests61PAMDAF
PAb Anti Mouse IgG-Eu cryptate - 5,000 tests61PAMKLA
PAb Anti Mouse IgG-Eu cryptate - 20,000 tests61PAMKLB
PAb Anti Mouse IgG-Tb cryptate - 5,000 tests61PAMTAA
PAb Anti Mouse IgG-Tb cryptate - 20,000 tests61PAMTAB
PAb Anti Mouse IgG-Tb cryptate - 1,000 tests61PAMTAF
PAb Anti Mouse IgG-XL665 - 5,000 tests61PAMXLA
PAb Anti Mouse IgG-XL665 - 20,000 tests61PAMXLB
PAb Anti Mouse IgG-XL665 - 1,000 tests61PAMXLF
PAb Anti Rabbit IgG-d2 - 5,000 tests61PARDAA
PAb Anti Rabbit IgG-d2 - 20,000 tests61PARDAB
PAb Anti Rabbit IgG-d2 - 1,000 tests61PARDAF
PAb Anti Rabbit IgG-Eu cryptate - 5,000 tests61PARKLA
PAb Anti Rabbit gG-Eu cryptate - 20,000 tests61PARKLB
PAb Anti Rabbit IgG-Tb cryptate - 5,000 tests61PARTAA
PAb Anti Rabbit IgG-Tb cryptate - 20,000 tests61PARTAB
PAb Anti Rabbit IgG-Tb cryptate - 1,000 tests61PARTAF
PAb Anti Rabbit IgG-XL665 - 5,000 tests61PARXLA
PAb Anti Rabbit IgG-XL666 - 20,000 tests61PARXLB
PAb Anti Rabbit IgG-XL665 - 1,000 tests61PARXLF
Protein A-Eu cryptate - 5,000 tests61PRAKLA
Protein A-Eu cryptate - 20,000 tests61PRAKLB
Protein A-XL665 - 5,000 tests61PRAXLA
Protein A-XL665 - 20,000 tests61PRAXLB

Companion products

DescriptionCat. noProduct insertMSDS
Streptavidin-XL665 - 5,000 tests610SAXLA
Streptavidin-XL665 - 20,000 tests610SAXLB
Streptavidin-XLent! - 5,000 tests611SAXLA
Streptavidin-XLent! - 20,000 tests611SAXLB
PPI - Terbium detection buffer - 200 mL61DB10RDF
PPI - Europium detection buffer - 200 mL61DB9RDF

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