Fluorescent Secondary Antibody Conjugates

Chromeo™ Dyes – Chromeo 488, Chromeo 494, Chromeo 505, Chromeo 546 and Chromeo 642 – are bright fluorescent labels that replace Alexa Fluor*, DyLight* or Cy*-dyes. They are compatible with most excitation sources including diode lasers, LEDs, tungsten lamps and xenon arc lamps. The dyes have been conjugated to high-quality secondary antibodies by an optimzed conjugation method, including subsequent purification from interfering substances.

Chromeo™ Dye Secondary Antibody Conjugate advantages

• High intensity
• Specifity under various fixation conditions
• Photostability – the combination with MAX Stain™ reagents provides optimal fluorescent stability for multiple exposures and increased exposure time
• Low background

Fluorescent Chromeo Dyes Secondary Antibody Conjugates enable sensitive and specific detection in immunofluorescence, high content screening, ELISA, FRET applications or flow cytometry. To ensure that you get the best results possible, we highly recommend that you use our secondary antibody conjugates together with our MAX Stain™ Immunofluorescence Tools, as those components have been formulated to optimize the performance of the Chromeo™ Dyes.

Dye	Absorption	Emission	Spectra	ε L/(mol-cm)	Stokes shift
Chromeo™ 488	488 nm	517 nm		73,000	29 nm
Chromeo™ 494	494 nm	628 nm		55,000	124 nm
Chromeo™ 505	505 nm	526 nm		70,000	21 nm
Chromeo™ 546	545 nm	561 nm		98,800	16 nm
Chromeo™ 642	642 nm	660 nm		180,000	18 nm
Table 1: Properties of Chromeo Dyes.

To receive more detailed information and application data about individual Chromeo dyes, simply "click" on the name or on the spectrum of your Chromeo dye of your choice in the table above.

Next to the spectral properties of the dye and the quality of the secondary antibody, the quality of of a fluorescent conjugate is influenced by the dye-to protein ratio, the conjugation method and its purity. All Active Motif Secondary Antibody Conjugates have been prepared by an optimized conjugation protocol making the fluorescent secondaries brighter and lowering the fluorescent background. Active Motif antibody conjugates have been tested in various applications including flow cytometry and fluorescent microscopy where they have shown to work with high efficiency and specificity under multiple fixation conditions.

Active Motif Fluorescent Secondary advantages

• Unrivaled fluorescent intensity
• Low background
• Limited photobleaching
• High Specificity
• Flexibility

Figure 1: Tubulin staining with Chromeo 488 secondary antibody.

U2OS cells stained with propidium iodide were incubated with rabbit anti-tubulin antibody. Chromeo 488 Goat anti-Rabbit IgG secondary antibody was then used at 1:1000.

Figure 2: EGFR staining with Chromeo 546 secondary antibody.

The Chromeo 546 Goat anti-Mouse IgG secondary antibody was used to stain EGFR in an urothel cell line. This data was generously provided by Dr. Brockhoff, Institut for Pathology, University Regensburg, Germany.

Contents & Storage

The Chromeo™ conjugates are supplied as 1 mg of antibody at a concentration of 2 mg/ml. The ATTO conjugates are supplied in 200 or 35 µl aliquots in PBS. For short-term storage, the conjugated antibody should be stored at 4°C protected from light. For longer-term storage, aliquot the antibody and store at -20°C. Avoid subjecting the antibody to repeated freeze-thaw cycles. These products are guaranteed for 6 months from the date of arrival.

To ensure photostability of Chromeo 488 under all experimental conditions, one aliquot of MAXfluor™ Mounting Medium for use in fluorescence microscopy is supplied with Chromeo 488 conjugates. This will provide optimal fluorescence stability and anti-fading during long-term storage of your mounted slides. MAXfluor Mounting medium should be stored at 4°C.

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