Background: Sperm preparation techniques aim at providing specimen for insemination with the highest quality spermatozoa independent of phenotypic and genomic integrity. Results of Assisted Reproductive Technologies (ART's) still do not exceed high percentages due to different factors, one of them is male factor infertility. Therefore the global companies concern on the techniques that used for activation of semen samples of infertile men. However, till now, there are many semen samples did not response to sperm preparation techniques leading to failure of sperm activation. New technological advancements in the field have altered current infertility management. Microfluidic Chamber is one of them, as it mimics the normal biophysical and biochemical environment and allows for more physiological sperm selection in ART's patients. Objective: The aim of this study was to compare the new sperm preparation technique by Microfluidic Sperm Sorting Chip or Microfluidic Chamber (MFC) with other techniques in asthenozoospermic Men. Patients and Methods: This study involved 30 males; the seminal fluid analysis from them was assessed and each semen sample was divided into 4 parts. The first part was prepared for sperm characterization and assessment of before or pre-activation, the second part using after or post-activation by Centrifugation technique, the third part using after activation by Simple layer, while the last part was using after activation by the new Microfluidic Chamber(MFC). Results: The MFC technique has the best results especially with the parameter of morphologically normal sperm in relation to other conventional techniques used in sperm preparation. Conclusion: Microfluidic Chamber is considered the best method for sperm preparation in comparison with other methods of sperm preparation in case of asthenozoospermic men.