Pre-analytical factors have a significant impact on the integrity of blood samples used for qualitative and quantitative protein profiling. Important factors are the type of the blood collection tube and the anticoagulant used. Only a few studies have been performed to assess these variables by comparing only serum and EDTA plasma collection tubes with some target proteins or peptides. In this study, we investigated the protein profile of blood samples collected in serum, EDTA-, heparin-, and citrate plasma tubes. Furthermore, we compared the depletion efficiency of 6 high abundant proteins, the detectable blood protein profile, the variance, and the coverage of the detectable protein sets. The largest differences were found between serum and plasma samples with respect to the peptide number and the occurrence of classical blood proteins. The heparin plasma evidenced a high number of detectable proteins, low global variance and a high similarity to EDTA- and citrate plasma and may therefore be also a useful test tube for blood protein profiling. In addition, a core set of blood proteins were described and the portions and compositions of sampling specific proteins were disclosed. Therefore, pre-analytical issues such as the sample collection method should be considered for protein profiling studies.