CD 45/67 immunobead preparation of human semen activates granulocytes

In human semen reactive oxygen species (ROS) produced by spermatozoa or leukocytes can impair spermatozoa functions. The objective of this study was to assess the effects of CD 45- and/or CD 67 immunobead preparation on the chemiluminescence (CL) of seminal plasma free ejaculate cells (= original cell suspension), as well as of the spermatozoa and leukocyte fractions. The original cell suspensions of 68 infertile and 8 fertile men were incubated with CD 45 or CD 67 immunobeads. After separation in a magnetic field the luminol chemiluminescence of the original cell suspensions, the spermatozoa and the leukocyte fractions were recorded on a luminometer. Spermatozoa fractions did not contain any leukocytes as no increase in CL-counts occurred after addition of FMLP. The number of peroxidase-positive cells (per 10(7) spermatozoa) correlated with the CL of the original cell suspensions (r = 0.7; P < 0.0001) as well as the CL of the spermatozoa and the leukocyte fractions after CD 45 or CD 67-preparation (r = 0.64; P < 0.0001). The CL of the spermatozoa and of the leukocyte fractions after CD 45 immunobead incubation were significantly correlated (r = 0.091; P < 0.0001). According to these data contaminating leukocytes could be eliminated by immunobead preparation. However, incubation of original cell suspensions with CD 45 or CD 67 immunobeads stimulated leukocytes to release soluble products resulting in elevated CL signals both in the leukocyte and the spermatozoa fractions. These effects have to be taken into account when using immunobeads for the preparation of human semen.