A homogeneous visualization and dual fluorescence signal analysis method and application based on multiple selective recognition reactions
A signal analysis method and fluorescence signal technology, which are applied in the field of homogeneous visualization and dual fluorescence signal analysis, and can solve problems such as difficult visualization and reading.
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Embodiment 1
[0039] This example provides a synthesis method of CdTe QDs.
[0040] First, 0.5 mmol CdCl 2 and 0.20 g of trisodium citrate were dissolved in 50 mL of water, and 52 μL of mercaptopropionic acid (MPA) was added to the above solution. Using NaOH solution, the pH of the above mixture solution was adjusted to 10.5;
[0041] Then, 0.1 mmol Na 2 TeO 3 and 50mg KBH 4 Add to the above solution, reflux for 1 hour, until the solution turns red, showing strong red fluorescence under UV lamp irradiation;
[0042] Finally, the CdTe QDs solution was purified by precipitation (using n-propanol) and centrifugation (11000 rpm, 30 min).
[0043] The MPA-CdTe QDs synthesized above were stored at 4°C before use.
Embodiment 2
[0045] The present embodiment provides the analysis steps of PPase, as follows:
[0046] (1) Add 77 μL Tris-HCl buffer (pH=7.4, 10 mM, 500 mM NaCl, 100 mM MgCl) to the centrifuge tube in sequence 2 ), 10 μL pyrophosphate (PPi, 17.5 mM) and 50 μL pyrophosphatase (PPase) with different concentrations, incubated at 37°C for 0.5 h to complete the hydrolysis reaction;
[0047] (2) Subsequently, 10 μL of CuCl was added to the centrifuge tube 2 solution (100 μM), continue to incubate at 37°C for 0.5h to complex copper ions and PPi;
[0048] (3) Second, add 2 μL Ce(NO) to the above centrifuge tube 3 ) 3 The solution (0.5mM) and 1μL of CdTe QDs stock solution were reacted at room temperature for 12min to complete the cation exchange reaction and the coordination complexation reaction;
[0049] (4) Transfer the above solution into a cuvette, use a fluorometer to test and record the data (excitation wavelength: 295 nm, emission wavelength range: 310 nm-800 nm).
Embodiment 3
[0051] This embodiment provides PPase analysis-excitation wavelength selectivity and selective identification phenomenon verification, as follows:
[0052] Considering that the optimal excitation wavelength for PPi-Ce CPNs is 295 nm, and the optimal excitation wavelength for CdTe QDs is 365 nm, the excitation wavelength of the dual fluorescence signal analysis system was selected first. like figure 2 As shown in A, when excited at 295 nm, both PPi-Ce CPNs and QDs have corresponding emission peak shapes, while when excited at 365 nm, PPi-Ce CPNs do not emit a peak at 348 nm, but show a peak shape at the excitation wavelength at 365 nm.
[0053] In summary, the final selectivity of 295 nm is the excitation wavelength of this system.
[0054] At the same time, it was further verified that different concentrations of PPi-Cu 2+ -The effect of PPi complexes on the fluorescence signal of QDs, and the effect of different PPi on Ce 3+ The effect of fluorescence signal. like figu...
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