1、 Signal strength
Ordinary indirect immunofluorescence(IF):
The signal strength is relatively weak, especially when detecting low abundance antigens, it may not be sensitive enough.
TSA multi-color immunofluorescence:
Due to the amplification effect of casein signal, the signal strength is significantly enhanced, allowing for the detection of lower abundance antigens and improving the sensitivity of detection.
2、 Principle
Ordinary indirect immunofluorescence(IF):
Firstly, add the primary antibody and combine with the antigen on the sample to form a complex.
Then add a fluorescently labeled secondary antibody, which binds to the primary antibody, and determine the presence and abundance of the test substance by detecting the fluorescence signal.
TSA multi-color immunofluorescence:
Based on casein signal amplification technology. Firstly, the primary antibody is incubated, and then the HRP labeled secondary antibody corresponding to the primary antibody is bound to the primary antibody. Then, fluorescein labeled casein is added, and under the catalysis of peroxidase, the fluorescein on casein covalently binds to the surrounding proteins of the secondary antibody, greatly amplifying the fluorescence signal.
This technology can perform multiple rounds of staining, and after each round of staining, the antibodies from the previous round are removed through specific processing, achieving high-resolution multi-color labeling of multiple targets on the same sample.
3、 Operational complexity
Ordinary indirect immunofluorescence(IF):
The operation is relatively simple, and the experimental process is relatively short.
TSA multi-color immunofluorescence:
The operation is relatively complex, requiring multiple rounds of staining and antibody removal steps, with high technical requirements and relatively long experimental time.
4、 Resolution
Ordinary indirect immunofluorescence(IF):
The resolution is relatively low, and for some finely structured tissues or cells, it may be difficult to clearly distinguish different antigen distributions.
TSA multi-color immunofluorescence:
• With high resolution, it can display the localization and distribution of different targets in cells or tissues more clearly, which is helpful for in-depth research on molecular interactions and spatial relationships within cells.
5、 Multi-color marking ability
Ordinary indirect immunofluorescence(IF):
Although multi-color marking is also possible, the number of multi-color markings is usually limited.
TSA multi-color immunofluorescence:
Multiple rounds of staining can be performed, and through precise control and signal amplification, more color labeling can be achieved, greatly improving the ability of multi-color labeling. Multiple different targets can be detected simultaneously, providing richer information for studying complex biological processes.