Protein A Acceptor Beads

Homogeneous Immuno Chemiluminescence Assay (HICA) is a homogeneous immunoassay method based on energy transfer between donor beads and acceptor beads at close proximity, resulting in luminescence.

Donor beads recognize Protein 1 (Tag1 label), while Acceptor beads recognize Protein 2 (Tag2 label). When Protein 1 binds to Protein 2, the distance between the beads becomes less than 200nm. Upon excitation at 680nm, the donor beads generate singlet oxygen, which diffuses to the acceptor beads. The acceptor beads then undergo a redox reaction, emitting light at 615nm. The signal intensity is directly proportional to the strength of the protein interaction.

This product features a simple operation process, no washing steps, fast speed, and high sensitivity, enabling the detection of weak interactions.

Specification	Fill Volume
250 μg	50 μL
5 mg	1 mL
25 mg	1 mL x 5

[Required Reagents]

Name	Cat. No.
Protein A Acceptor Beads	UA086091
Streptavidin Donor Beads	UA086104
Universal Buffer 1	UA086113

[Assay Protocol for Reference]

Assay Protocol	Protocol 1 (37°C Rapid Detection)	Protocol 2 (Room Temperature Detection)
Step 1:	4μL Tag1-M1 +4μL Tag2-M2+ 6μL Acceptor Beads, Protect from light / Green light	4μL Tag1-M1 +4μL Tag2-M2+ 6μL Acceptor Beads, Protect from light / Green light
Incubation	Incubate at 37°C with shaking for 20 minutes, Protect from light / Green light	Incubate at room temperature for 60 minutes, Protect from light / Green light
Step 2:	Add 6μL Donor Beads, Protect from light / Green light	Add 6μL Donor Beads, Protect from light / Green light
Incubation	Incubate at 37°C with shaking for 10 minutes, Protect from light / Green light	Incubate at room temperature for 30 minutes, Protect from light / Green light
Read	Instrument reading	Instrument reading

[Performance Validation]

•Sample Preparation:

Pre-dilute biotinylated rabbit IgG (Bio-rIgG) to 15 μg/mL (100 nM) using Universal Buffer 1 as a stock solution, then perform serial dilutions according to the following scheme:

ID	Final Concentration (nM)	Universal Buffer 1 Volume (μL)	Volume of Higher Concentration Added (μL)
C12	1.0E+01	210	90μL Stock solution
C11	3.0E+00	210	90μL C12
C10	1.0E+00	180	90μL C11
C9	3.0E-01	210	90μL C10
C8	1.0E-01	180	90μL C9
C7	3.0E-02	210	90μL C8
C6	1.0E-02	180	90μL C7
C5	3.0E-03	210	90μL C6
C4	1.0E-03	180	90μL C5
C3	3.0E-04	210	90μL C4
C2	1.0E-04	180	90μL C3
C1	0	180	/

•Detection Reagent Preparation:

Name	Preparation Concentration	Diluent
Protein A Acceptor Beads	25 μg/mL	Universal Buffer 1
Streptavidin Donor Beads	25 μg/mL	Universal Buffer 1

•Results for 37°C Incubation Mode:

Maximum Signal: 7151971

Minimum Signal: 866

EC50= 0.195 nM

•Results for Room Temperature Incubation Mode:

Maximum Signal: 3031800

Minimum Signal: 389

EC50= 0.176 nM

1. This experiment is light-sensitive; ensure all operations are performed in the dark. It is recommended to conduct preparation, sample addition, and incubation steps under green light (illuminance below 100 LUX). 2. This product is compatible with multifunctional microplate readers equipped with Alpha detection modules. 3. Vortex thoroughly before use, or briefly centrifuge (2000×g, 5–10 seconds) to ensure complete usage. 4. It is recommended to use the company's配套 diluent for reagent preparation and sample dilution. If additional components are required, they can be directly added to this diluent. 5. To ensure comparability of experimental data across different batches, strictly control incubation temperature and time. 6. Avoid generating bubbles during sample addition.