Human luteinizing hormone (LH) is an important analyte for doping control analysis since it increases the athletic performance. However, traditional methods to detect LH have few disadvantages, such as long analysis duration, waste disposal problem and sample matrix effect. Addressing these problems, surface enhanced Raman spectroscopy based LH analysis using a passive microfluidic chip was developed and optimized. Antibody modified magnetic gold nanoparticles captured the LH and then, 4-aminothiophenol (4-ATP) labeled nanoparticles formed the sandwich immunoassay structure. The complex and the other reactions occurred in different chambers of the chip. The SERS signals of 4-ATP were recorded from the chamber and the system was shown to detect 0.036 IU L-1 in serum samples. The performance of the immunoassay was compared to all other methods and the proposed assay was the fastest analysis of LH without any problems associated with the sensitivity. The shorter analysis time was recorded because the chip enables the control of all reactions in one place and there was no requirement of a specialized laboratory. (c) 2018 Elsevier B.V. All rights reserved.