When designing multicolor flow cytometry experiments, the choice of fluorophore is important. However, we found that the name of the labeled fluorescence of the flow cytometry instrument or the fluorescein written in the detection instructions provided by the manufacturer are not

When designing multi-color flow cytometry experiments, the choice of fluorescein is very important. However, we found that the name of the labeled fluorescence of the flow cytometry instrument or the fluorescein written in the detection instructions provided by the manufacturer are not so comprehensive. Due to factors such as different brands of raw materials, patent restrictions, and fluorescein research and development, we often encounter many unfamiliar fluorescein names. So how do we judge whether our own flow cytometer can detect these fluorescences?

Let’s first understand the characteristics of fluorescein, two characteristic spectra (nm):

① Excitation spectrum (Excitation, Ex): refers to the light within a certain wavelength range that can specifically excite a certain fluorescein, also called Absorption spectrum . The maximum absorption wavelength, which is the absorption peak, is referred to as Ex-Max.

② Emission spectrum (Emission, Em): refers to the fluorescence within a certain wavelength range caused by fluorescein emitted by excitation light of a certain wavelength. The maximum emission wavelength, that is, the emission peak, referred to as Em-Max.

Let’s take a look at the conventional flow cytometer and the principle of detecting fluorescein:

The optical system of the flow cytometer is equipped with different lasers and filters , which are used to excite and collect different fluorescein signals. .

① Laser: used to provide excitation energy to fluorescein. Common excitation lights include purple Violet (405nm), blue Blue (488nm), green (561nm), red (633nm), and high-end instruments also include ultraviolet light (345nm) to excite fluorescein.

② Filter: It can be divided into long pass, short pass and band pass. It is used to collect different ranges of emission wavelengths, that is, it can detect the emission signals of different fluoresceins. The setting of the filter is mainly for the occurrence of spectra. The emission peak is set.

Therefore, when we usually talk about color matching, we need to provide the fluorescence configuration of the flow cytometer. It is necessary to have two pieces of information about the specific exciter and filter to determine the specific fluorescence channels that can be detected.

The picture below is an example, which shows the filter installed under the 405nm laser and the fluorescein that can be detected:

However, we found that the fluorescein commonly used under the 405nm laser, such as the BV series fluorescein, is not available on the flow cytometer Please note, can I still use BV dyes when color matching? Here is a confirmation method for you:

We can use the "Fluorescence Spectrum Viewer" on the BD official website (https://www.bdbiosciences.com/zh-cn/resources/bd-spectrum-viewer) to check Its fluorescein excitation wavelength and filter information.

As shown in the picture above, as long as the excitation wavelength and emission wavelength of fluorescein are similar, can be excited by the exciter, and can be within the receiving wavelength range of the filter, it can be detected. For example, BV421 can actually be detected using the V450 or Pacific Blue channel.

If you think this is more complicated, or if the mobile phone is more convenient, you can also query it through the fluorescence spectrum viewer in the WeChat public account of our " streaming expert ".

Don’t miss out on a lot of bright, cost-effective fluorescein just because the name is unfamiliar to you!

BD Sirigen series of patented fluorophores: BB series, BUV series, BV series, APC-R700, R718, brighter, more stable and more accurate.