Patent ID: 7875442

Claim:
A process for real-time PCR of multiple target nucleotide sequences comprising the steps of: a) providing a closed reaction chamber containing a PCR solution and covalently immobilized unlabeled capture molecules wherein said capture molecules are immobilized in specifically localized areas of a solid support in the form of a micro-array of at least 4 different capture molecules per cm 2 , wherein the PCR solution comprises multiple target nucleotide sequences, primers specific to said target nucleotide sequences, and a fluorescent label and wherein the micro-array is in intermittent contact with the PCR solution; b) carrying out the following steps without opening the closed reaction chamber: (i) conducting at least one PCR cycle to form amplicons of the target nucleotide sequence wherein said amplicons arc formed with fluorescent label obtained using either labeled primer or by enzymatic incorporation of labeled nucleotide; (ii) hybridizing the amplicons to immobilized unlabeled capture molecules; and (iii) detecting the hybridized amplicons wherein the PCR solution contained in the chamber is moved away from the capture molecules during said detection; (iv) repeating steps (i)-(iii) at least once c) performing identification and/or quantification of the target nucleotide sequence corresponding to said capture molecules during the amplification cycles without washing.