The study of biomarkers in the cerebrospinal fluid (CSF) of patients with mild cognitive impairment (MCI) is a technique used with increasing frequency in the early diagnosis of Alzheimers disease (AD). Our objectiv was to gain an own experience while evaluating the reliability, sensitivity, and reproducibility of this technique in Spanish patients. Thirty-seven patients with MCI and twenty-four control subjects were studied by means of AD biomarker analysis in CSF. xMAP Luminex and INNO-BIA Alzbio3 reagents of Innogenetics were used. The study variables assessed were levels of Aβ(1-42), T-tau and P-tau(181p) proteins as well as the ratios of T-tau/Aβ(1-42) and P-tau(181p)/Aβ(1-42). Samples from nineteen patients were examined twice. Intra-class correlation coefficients for the three biomarkers used showed values higher than 0.95. We observed significant differences between the control group and the MCI groups. In the 6 months following lumbar puncture (LP), eleven (29%) patients with MCI developed AD. These patients showed significant lower levels in Aβ(1-42) protein (276.35 ± 78 vs. 367.13 ± 123.49, P < 0.03) and higher ratios (T-tau/Aβ(1-42) [0.38 ± 0.2 vs. 0.22 ± 0.14, P < 0.01] and P-tau(181p)/Aβ(1-42) [0.27 ± 0.13 vs. 0.16 ± 0.1, P < 0.008]) to those in the same group who remained stable. We obtained similar results to those in the most recent reliable literature with our ROC curves, especially with our P-tau(181p) values and T-tau/Aβ(1-42) ratio in order to differentiate between control and AD groups. Our experience showed that the analysis of CSF-AD biomarkers in patients with MCI is reliable, sensitive and reproducible. In our knowledge, this is the first experience in Spanish patients.