Products

Amine-Activated Terbium Fluorescence Magnetic Beads

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2.5 μm BcMag™ Aldehyde-Activated Terbium Fluorescence Magnetic Beads
Cat. No.  GE101

Unit Size  15 mg
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2.5 μm BcMag™ Aldehyde-Activated Terbium Fluorescence Magnetic Beads
Cat. No.  GE102

Unit Size  30 mg
Order
5 μm BcMag™ Aldehyde-Activated Terbium Fluorescence Magnetic Beads
Cat. No.  GE103

Unit Size  15 mg
Order
5 μm BcMag™ Aldehyde-Activated Terbium Fluorescence Magnetic Beads
Cat. No.  GE104

Unit Size  30 mg
Order
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Cat. No.

Product Name

Unit Size

Order

GE101

2.5 μm BcMag™ Amine-Activated Terbium Fluorescence Magnetic Beads

15 mg

GE102

2.5 μm BcMag™ Amine-Activated Terbium Fluorescence Magnetic Beads

30 mg

GE103

5 μm BcMag™ Amine-Activated Terbium Fluorescence Magnetic Beads

15 mg

GE104

5 μm BcMag™ Amine-Activated Terbium Fluorescence Magnetic Beads

30 mg

Fluorescence Dye Properties

Fluorophore

Fluorescence
Color

Excitation
(nm)

Emission
(nm)

Fluorescence lifetime
(Ԏ) (µsec)

Stokes shifts
(nm)

Selection of
Emission Filter

Terbium (Tb3+)

Green

320

545

1050

220

545/40

Fluorescence Dye Properties

Fluorophore

Terbium (Tb3+)

Fluorescence
color

Green

Excitation
(nm)

320

Emission
(nm)

545

Fluorescence lifetime (Ԏ) (µsec)

1050

Stokes shifts
(nm)

220

Selection of
Emission Filter

545/40

Specification

Bead Size

2.5μm diameter; 5μm diameter

Number of Beads

~ 1.47 x 108 beads/mg (2.5μm beads)

~ 5 x 107 beads /mg (5μm beads)

Stability

Short Term (<1 hour): pH 3-11; Long-Term: pH 4-10

Temperature: 4°C -140°C; Most organic solvents

Magnetization

~ 40-45 EMU/g

Type of Magnetization

Superparamagnetic

Formulation

Lyophilized Powder

Functional Group Density

2.5μm Magnetic Beads

~240 μmole / g of Beads

5μm Magnetic Beads

~200 μmole / g of Beads

Storage

Store at 4°C upon receipt. Do not freeze.

Description

BcMag™ Amine-activated Terbium Fluorescence Magnetic Beads are Time-Resolved Fluorescence (TRF) magnetic microspheres coated with a high density of primary amine functional groups on the surface. The microspheres combine the benefits of the preactivated amine active group, time-resolved Fluorescence dyes, and magnetic characteristics to perform very sensitive assays. The beads are manufactured using nanometer-scale superparamagnetic iron oxide and europium metal as core and entirely encapsulated by a high-purity silica shell, ensuring no leaching problems with the iron oxide and europium metal.

Amine beads coupling reaction
BcMag™ Terbium Fluorescence Magnetic Beads

The carboxyl group is found in a wide range of biological substances. Peptides and proteins contain carboxyls (-COOH) at the C-terminus of each polypeptide chain, as well as aspartic acid (Asp, D) and glutamic acid side chains (Glu, E). Carboxyls, like primary amines, are typically found on the surface of protein structures.

Carboxylic acids can be employed to immobilize biological molecules by applying a carbodiimide-mediated process. Although no activated support has a reactive group that is spontaneously reactive with carboxylates, chromatographic supports containing amines (or hydrazides) can be utilized to create amide bonds with carboxylates activated using the water-soluble carbodiimide crosslinker EDC.

Although conventional fluorophores have been widely used over the past decades, they still suffer from either one or several limitations in terms of applicability and efficiency:

1.

Narrow excitation bands cause higher background signals.

2.

Smaller Stokes shift often produces self-quenching.

3.

Fluorescence is sensitive to environmental factors such as metallic ion concentration, pH, temperature, and solvent polarity.

4.

Fluorescence intensity is not high enough for detecting a single biomolecular.

5.

Fluorescence intermittency (blinking) affects some processes of molecule detection.

6.

Easily aggregated because of hydrophobicity.

BcMag™ TR-FRET (Time-Resolved FRET) Assay

BcMag™ TR-FRET Assay, in contrast to typical FRET (Förster Resonance Energy Transfer) assays, uses time-resolved Fluorescence magnetic beads (BcMag™ TR-Magnetic Beads) as the donor fluorophore. The donor and acceptor can be two proteins, two DNA strands, an antigen, an antibody, or a ligand and its receptor. After a reasonable time delay (usually 50 to 100 s), a signal is generated by fluorescence resonance energy transfer between a donor and an acceptor molecule when they are close and monitored in a time-resolved way. In BcMag™ TR-FRET Assay, a trace amount of analytes can be easily enriched from the complex by TR-Magnetic Beads, resulting in higher sensitivity. This assay practically eliminates all fluorescence backgrounds caused by the sample and plastic microplate, as well as by direct acceptor excitation. As a result, the signal-to-noise ratios of the BcMag™ TR-FRET Assay are very high, and the background is quite low. Furthermore, the assay does not need washing steps. BcMag™ TR-FRET Assay offers substantial advantages to bioassays in high throughput screening, such as assay flexibility, dependability, increased assay sensitivity, higher throughput, and fewer false positive/false negative results.

Terbium cryptate fluorophore is an efficient Fluorescence label due to its distinct specific properties. It is excited at 320nm and emits green fluorescence at 545nm, with a long fluorescence lifetime (1050 µsec) and large stokes shifts (220 nm). By taking advantage of these properties, time-resolved fluorescence measurement can dramatically reduce the fluorescence background from the sample and increase the signal-to-noise ratio to offer detectability better than one order of magnitude than conventional Fluorescence dyes. BcMag™ Amine-activated Terbium Fluorescence Magnetic Beads are excellent donors used in TR-FRET assays.

Workflow of TR-FRET Magnetic Beads Assay

1.

Mix the antibody-conjugated donor beads with the cell lysates and incubate them with continuous rotation for a sufficient time. The beads remain suspended in the sample solution during mixing, allowing the target analytes to bind to the donor beads.

2.

After incubation, the beads are collected and separated from the sample using a magnet rack.

3.

Add the antibody-conjugated acceptor and incubate them with continuous rotation for a sufficient time.

4.

Analysis of numerous microplate readers supports TR-FRET measurements.

Workflow of TR-FRET Magnetic Beads Assay

Advantages and Benefits

1.

Perform a double function simultaneously on the same beads: The magnetic beads combine separation/preconcentration and detect analytes, allowing quick, simple, robust, and high-throughput analytes of trace amounts from complex biological samples on the same beads.

2.

Ultra sensitive. Lower detection limits of 10 pg/mL versus typical fluorometric detection limits of 100 pg/mL.

3.

Extremely photostable and highly resistant to photobleaching. All the lanthanide chelate or cryptate molecules and iron oxide are entirely encapsulated inside each bead instead of merely on the bead’s surface. The protective environment prevents iron oxide and dye from leaching into aqueous media, which makes the beads less sensitive to external conditions such as solvent, temperature, pH, etc.

4.

Very high Fluorescence intensity. Because a single bead has a large concentration of lanthanide chelate with a high quantum yield ranging from 40 to 90%, the beads show excellent fluorescence intensity, which increases test sensitivity without signal amplification. Such bright beads are also perfect for donors’ use in time-resolved FRET assays.

5.

Lanthanide chelate or cryptate has large Stokes shifts (>250 nm), narrow emission bands (-10 nm bandwidth), and long fluorescence lifetime (μs), which dramatically reduces background and increases the signal-to-noise ratio.

6.

Most bioprocess ELISA assays can be converted to an HTRF assay.

7.

No washing step is involved in the assays.

8.

Have a hydrophilic silica surface grafted by different functional groups with linkers of variable lengths, allowing efficient conjugation of various ligands such as peptides, proteins, antibodies, small molecules, carbohydrates, aptamers, DNA/RNA, etc.

9.

Due to the microsphere’s magnetic properties, the Fluorescence magnetic beads are suitable for high-throughput automation.

Conjugation Protocol

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