Method and kit for simultaneous detection of multi target molecules using magnetic bead-aptamer conjugate

A method for simultaneous detection of multi target molecules using a magnetic bead-aptamer complex, comprising:

• (1) incubation
• incubating the magnetic bead-aptamer complex with a target molecule-containing sample to form a magnetic bead-aptamer-target molecule complex, wherein the aptamer are a group of aptamers screened by SELEX and specific to the target molecule-containing sample;
• and further comprising the step of:
• preparing the magnetic bead-aptamer-target molecule complex by incubating 100 μL of the target molecule-containing sample with 50 μL of the magnetic bead-aptamer complex at 37° C. for 30 min, washing the reaction mixture once with 3×SSC;
• and washing the reaction mixture three times with 0.4 mM binding buffer each for a volume of 20 times the volume of the reaction mixture followed by magnetic separation to obtain the magnetic bead-aptamer-target molecule complex:
• (2) acid elution
• adding an elution buffer to the magnetic bead-aptamer-target molecule complex under shaking to collect an eluate to which Tris-HCl containing a standard peptide is added to neutralize the eluate for detection of the target molecules, wherein the acid elution comprises the steps of:
• adding 0.5 mL of the elution buffer to the magnetic bead-aptamer-target molecule complex obtained in step (1) followed by shaking for 1 min to collect the eluate;
• and immediately neutralizing the eluate with 250 μL of 1 M Tris-HCl (pH 8.0):
• repeating the elution and neutralization twice;
• and combining the neutralized eluates to obtain a target molecule solution for mass spectrometry;
• (3) subjecting the neutralized eluate to mass spectrometry for qualitative and quantitative detection of the target molecules;
• and
• (4) analyzing data from the mass spectrometry on a data analytics platform to obtain the detection results.