MAO Probe 1 

MAO Probe 1 is a reactive two-photon fluorescent probe capable of crossing the blood-brain barrier, which is used to detect the activity of monoamine oxidases (MAO-A/MAO-B), with K_m values of 70 μM (MAO-A) and 75 μM (MAO-B), respectively. IBC 2 (benzo[g]imino-coumarin 2), the enzymatic product of MAO Probe 1, can further specifically bind to and image Aβ plaques, enabling in vivo two-photon co-monitoring of MAO activity and Aβ plaques. MAO Probe 1 can be used in Alzheimer's disease research (IBC 2: Ex/Em = 850 nm/570-620 nm)^[1].

MAO Probe 1 functions as a substrate for purified MAO-A and MAO-B, with measured K_m values of 70 μM (MAO-A) and 75 μM (MAO-B), kcat values of 71 min⁻¹ (MAO-A) and 53 min⁻¹ (MAO-B), and kobs values of 5.8 s⁻¹ (MAO-A) and 6.4 s⁻¹ (MAO-B) in pH 7.4 HEPES buffer at 25 °C^[1].
MAO Probe 1 (10 μM; 1 h) does not exhibit a fluorescence response when incubated with aggregated Aβ_1-42 plaques at 25 °C in PBS buffer (pH 7.4)^[1].

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

MAO Probe 1 (10 mg/kg; i.p.; single administration) enables synchronous in vivo monitoring of MAO activity and Aβ plaque burden in 5XFAD mice. Its fluorescence intensity (reflecting MAO activity and AD progression) exhibits three distinct stage-specific changes with increasing age: a slow initiation stage before 3 months of age, a rapid progression stage from 4 to 9 months of age, and a plateau stage after 9 months of age^[1].
MAO Probe 1 (10 mg/kg; i.p.; single administration) shows only negligible fluorescent signals in 9-month-old healthy wild-type mice, confirming its ability to distinguish healthy mice from Alzheimer's disease mice^[1].

Guide (The following is our recommended experimental protocol. This protocol serves as a reference guide only. Specific operations should be adjusted according to your actual needs.)
1. Preparation of Probe 1/IBC 2 stock solution
Dissolve in DMSO. Prepare a high concentration stock solution (usually 10-20 mM), protected from light, stored at -20°C.
Working solution is diluted with PBS/aCSF to the desired concentration (10 μM for tissue staining; according to body weight for in vivo).
2. In vivo dual-photon co-monitoring of MAO activity + Aβ plaques
2.1 Animal model: 2-11 month old 5XFAD transgenic mice (Tg6799) and age-matched WT controls
2.2 Probe injection: Intraperitoneal injection of Probe 1, dose 10 mg/kg body weight
Example: 20 g mouse → 200 μg Probe 1, dissolved in an appropriate solvent (commonly 5% DMSO + PBS or normal saline for dissolution)
After injection, incubate for 2 hours to allow the probe to cross the blood-brain barrier and be converted to IBC 2 by MAO
2.3 Surgical preparation (thin skull window / craniotomy)
2.3.1 Anesthesia: Zoletil 50 + Rompun (1.2 mL/kg, im), fixed on a 37°C heating plate
2.3.2 Disinfect the scalp, select a region 0.5-1 mm from the anterior fontanelle, and grind the skull to ~30 μm
2.3.3 Attach a circular cover glass + dental cement for fixation
2.4 Dual-photon imaging
Excitation wavelength: 850 nm
Emission and acquisition: 570-620 nm (red channel, IBC 2); can simultaneously collect Dextran-Texas Red (blood vessels, ~590-650 nm)
2.5 Control group (optional):
1 week before, administer Selegiline (HY-14198) (MAO-B inhibitor, 10 mg/150 mL ad libitum) in the drinking water, and proceed with the same steps. Expected signal significantly reduced.
3. Ex vivo brain tissue section Aβ plaque staining
3.1 Exsanguinate 5XFAD mice, quickly remove the brain
3.2 Use a vibrating slicer (Leica) to make horizontal slices of the hippocampal region (on ice, aCSF continuously perfused with 95% O₂/5% CO₂)
Incubate in aCSF containing IBC 2 (10 μM) + MeO-X04 (10 μM, Aβ reference dye) for 45 minutes (33°C)
3.3 Wash with aCSF 2-3 times.
3.4 TPM imaging: Ex = 850 nm, Em = 570-620 nm.
Note: In vitro experiments can also use IBC 2 to directly stain plaques for verification; in vivo, only use Probe 1 (in situ generated IBC 2 by MAO).

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

320.39

C₁₉H₂₀N₄O

1386860-79-2

NCCCOC1=CC2=CC(N(C)C)=CC=C2C=C1/C=C(C#N)/C#N

Room temperature in continental US; may vary elsewhere.

Please store the product under the recommended conditions in the Certificate of Analysis.

• [1]. Kim D, et al. Close Correlation of Monoamine Oxidase Activity with Progress of Alzheimer's Disease in Mice, Observed by in Vivo Two-Photon Imaging. ACS Cent Sci. 2016;2(12):967-975.