TrailBlazer™ Tag and TrailBlazer StarBright™ Dye Label Kits in Veterinary Studies

Expanding Panels with Cross-Reactive and TrailBlazer Kit-Labeled Antibodies

Although Bio-Rad has the largest range of feline antibodies, present fluorophore options restrict feline panel sizes, limiting the number of cell populations that can be identified. One of several possible 4-color panels is shown in this example (Table 1 and Figure 1), where CD4+ T cells (T helper cells) and their activation status can be identified in feline blood.

Table 1. Reagents used in the 4-color feline panel.

Fig. 1. 4-color feline panel. Red cell lysed feline blood was stained with a 4-color panel (Table 1) and acquired on a ZE5 Cell Analyzer. Single cell gate not shown.

To enable identification of B cells and monocytes, validated cross-reactive antibodies against CD3, CD14, and CD45RA were introduced, expanding the panel to 7-colors (Table 2 and Figure 2).

Table 2. Reagents used in the 7-color feline panel, including cross-reactive antibodies.

* Cross-reactive antibodies.

Fig. 2. 7-color feline panel including cross-reactive antibodies. Red cell lysed feline blood was stained with the panel shown in Table 2 and acquired on a ZE5 Cell Analyzer. Single cell gate not shown.

To enable identification of all major blood immune cell populations in this panel, including T helper cells, cytotoxic T cells, granulocytes, monocytes, and B cells, the panel was further expanded with TrailBlazer StarBright Dye Kit-labeled antibodies against CD8, CD18, and CD21, creating a larger 10-color panel (Table 3 and Figure 3).

Table 3. Reagents used in the 10-color feline panel. StarBright Dye TrailBlazer Kit-labeled antibodies were generated by using a TrailBlazer Tag Kit (12020038) and the indicated TrailBlazer StarBright Dye Label Kit.

* Cross-reactive antibodies. 

Fig. 3. 10-color feline panel incorporating cross-reactive antibodies and StarBright Dye TrailBlazer Kit-labeled antibodies. Red cell lysed feline blood was stained with a panel (Table 3) and acquired on a ZE5 Cell Analyzer. Single cell gate not shown. TrailBlazer Kit-labeled antibodies are highlighted in green.

Identifying a Low Antigen Density T Cell Marker with TrailBlazer Kit-Labeled Antibodies

TrailBlazer Kits can be used to label antibodies against a wide range of targets, including low-density markers and markers expressed on small cell populations. In pigs, CD27 is an important marker for assessing T cell memory status, but limited fluorophore availability and low antigen density—which require a bright fluorophore—can make it challenging to include in larger panels. This challenge was addressed by conjugating the CD27 antibody to the bright StarBright Violet 670 fluorophore (brightness 5/5). TrailBlazer Kits also enabled CD44, a pan T cell marker, to be added to the panel using the slightly less bright but spectrally narrow StarBright Violet 515 dye (brightness 4/5).

The data (Figure 4) were consistent with published literature, and clearly identified distinct naive, central memory (CM), and effector memory (EM) T cell populations. The small increase in expression of CD11a as cells differentiated from naive to EM cells within the CD8 memory populations confirmed the validity of these populations (Álvarez et al., 2025).

Table 4. Reagents used in the porcine 9-color T cell panel. StarBright Dye TrailBlazer Kit-labeled antibodies were generated by using a TrailBlazer Tag Kit (12020038) and the indicated TrailBlazer StarBright Dye Label Kit.

Fig. 4. Porcine 9-color T cell panel. Red cell lysed porcine blood was stained with an antibody panel shown in Table 4 and acquired on a ZE5 Cell Analyzer. TrailBlazer Kit-labeled antibodies are highlighted in green.

Large Immunophenotyping Panel—Multiplexing TrailBlazer Kit Conjugates

Beyond the cell type-specific panel, TrailBlazer Kits enabled a previously published 10-color general immunophenotyping panel to be expanded into a larger 19-color panel for high-dimensional immunophenotyping. This expanded panel enables deeper characterization of naive, central memory (CM), and effector memory (EM) T cells, regulatory T cells (T regs), B cell populations, and γδ T cells. Populations that previously required three separate panels to identify (Clark et al., 2019) can now be assessed in a single panel.

Table 5. Reagents used in the expanded 19-color porcine panel. StarBright Dye TrailBlazer Kit-labeled antibodies were generated by using a TrailBlazer Tag Kit (12020038) and the indicated TrailBlazer StarBright Dye Label Kit.

* Cross-reactive antibody. Antibodies from Bio-Rad unless indicated.

Fig. 5. Expanded 19-color porcine panel incorporating cross-reactive antibodies and StarBright Dye TrailBlazer Kit-labeled antibodies. Red cell lysed porcine blood was stained with an antibody panel shown in Table 5 and acquired on a ZE5 Cell Analyzer. TrailBlazer Kit-labeled antibodies are highlighted in green.

TrailBlazer Tag and StarBright Dye Label Kits allow purified antibodies to be conjugated to one of 32 StarBright Dyes, increasing fluorophore choice and enabling larger immunophenotyping panels.

Yes. Cross-reactive antibodies are commonly used when species-specific reagents are unavailable. Combining these antibodies with TrailBlazer-conjugated antibodies can expand panel size substantially.

Any species where purified antibodies are available, including feline, porcine, canine, bovine, and other veterinary research models.

Directly conjugated antibodies reduce workflow complexity and eliminate unwanted binding caused by secondary antibodies.