Elab Fluor^® 647 Anti-Mouse IL-17A Antibody[TC11-18H10.1] (E-AB-F1199M)

Q1:When do flow antibody samples use sterile consumables?

Subsequent experiments requiring cell culture procedures require the use of sterile consumables, such as intracellular factor testing or flow sorting experiments.

Q2:What are the precautions for detecting intracellular indicators in tissue samples?

① All sample tubes should be fixed and broken as in the experimental group, such as blank control, single standard group, fmo-iso; ② The fixed breaking time of each group should not be different for too long. If the sample size is large, it is recommended to operate in batches; ③ For tumor tissue samples, due to the small number of cells for detection, it is necessary to consider increasing the amount of antibodies and the number of computer time should be at least 1 million; ④ It is recommended to add 75um micron aperture filter for secondary filtration to improve the purity and content of immune cells; (5) Due to the longer stimulation blocking time, it is necessary to pay attention to aseptic operation before the end of the stimulation blocking time.

Q3:Can Th and Treg cells (requiring cell membrane vs. nuclear membrane permeabilization) be detected in a single tube?

Theoretically possible, but separate permeabilization buffer are recommended for optimal results: Nuclear membrane permeabilization buffer (stronger, ideal for nuclear targets like FOXP3) may damage cell membrane epitopes. Cell membrane permeabilization buffer (milder, better for cytoplasmic/membrane targets like cytokines).

Q4:Why centrifuge before use?

During the transportation of antibodies, antibodies will stick to the tube wall or cap due to turbulence. So after receiving the antibodies, moderate centrifugation will collect the antibodies on the tube wall or cap to the bottom of the tube to avoid the loss of antibodies.

Q5:What is the difference between the test-package and the weight-package of flow antibody products?

The usage of test-package antibodies is well designed and verified there is no need for an extra dilution before use. The weight-package antibody has a higher concentration, and requires a titration process for a suitable usage amount.

Q6:What is the concentration of primary antibody?

Our antibodies are concentrated, and the final concentration of the antibodies is not only related to the concentration of the antibodies themselves, but also to the abundance of the target protein in the sample. Therefore, the recommended dilution ratio for the antibodies provided in our instructions is a range, and the final dilution ratio needs to be determined through pre-experiments.You can also consult our technical team to get a recommendation of a suitable dilution ratio for your experimental conditions.

Q7:What does Isotype Control do? How to choose a suitable isotype control antibody?

Isotype control antibodies are used as the basis for determining negative and positive cells. It is necessart as a gating helper especially for the indicators with low expression or continuous expression.
The Isotype control was purified from the serum of non-immunized animals, it should be the same species source, same immunoglobulin and subtype, same fluorescein label, same dose and concentration as the stained monoclonal antibody.

Q8:What does IHC mean on the datasheet? Is it a frozen tissue section? How is it different from IHC-F?

IHC is an immunohistochemical experiment, and the sections used generally include IHC-P (paraffin tissue section) and IHC-F (frozen tissue section).

Q9:What auxiliary reagents are needed for staining?

For samples with erythrocyte, ACK lysis buffer (E-CK-A105) is needed;
For cells that are rich in Fc receptors, such as macrophages, Fc receptors blocking is necessary before staining with flow Antibody to reduce the non-specific signal. At present, we can provide human and mouse blocking agent, E-AB-F1236A Purified Anti-Human CD16 Antibody[3G8]. E-AB-F0997A Purified Anti-Mouse CD16/32 Antibody[2.4G2].
Cell staining buffer (E-CK-A107) is required in the process of cell staining.
For the detection of intracellular indexes, a fixation & permeabilization kit is needed (E-CK-A109) ; and for the detection of intranuclear indexes, a specific staining kit (E-CK-A108) is required.
Dead cell dyes are also used for flow cytometry of tissue samples such as tumors.

Q10:What are the requirements for centrifuge usage when preparing samples?

The centrifugal force of the centrifuge does not exceed 300g, the centrifugal speed does not exceed 3, and the centrifugal speed does not exceed 1, so as to avoid cell damage.