Examination and processing of human semen - libdoc.who.int ...

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v Chapter 3 Optional procedures 115 3.1 Indices of multiple sperm defects 115 3.1.1 Calculation of indices of multiple morphological defects 115 3.1.2 Worked example 116 3.2 Panleukocyte (CD45) immunocytochemical staining 117 3.2.1 Principle 117 3.2.2 Reagents 118 3.2.3 Procedure 118 3.3 Interaction between spermatozoa and cervical mucus 122 3.3.1 In-vivo (postcoital) test 122 3.3.2 In-vitro tests 125 3.3.3 In-vitro simplifi ed slide test 126 3.3.4 Capillary tube test 127 3.4 Biochemical assays for accessory sex organ function 130 3.4.1 Measurement of zinc in seminal plasma 130 3.4.2 Measurement of fructose in seminal plasma 132 3.4.3 Measurement of neutral -glucosidase in seminal plasma 134 3.5 Computer-aided sperm analysis 136 3.5.1 Introduction 136 3.5.2 Use of CASA to assess sperm motility 137 3.5.3 Use of CASA to estimate sperm concentration 140 3.5.4 Computer-aided sperm morphometric assessment 140 Chapter 4 Research procedures 142 4.1 Reactive oxygen species 142 4.1.1 Introduction 142 4.1.2 Measurement of reactive oxygen species generated by sperm suspensions 143 4.2 Human sperm–oocyte interaction tests 146 4.3 Human zona pellucida binding tests 146 4.4 Assessment of the acrosome reaction 147 4.4.1 Procedure for the fluorescence assessment of acrosomal status 147 4.4.2 Induced acrosome reaction assay 150 4.5 Zona-free hamster oocyte penetration test 152 4.5.1 Protocol 152 4.6 Assessment of sperm chromatin 157 PART II. SPERM PREPARATION Chapter 5 Sperm preparation techniques 161 5.1 Introduction 161 5.1.1 When spermatozoa may need to be separated from seminal plasma 161 5.1.2 Choice of method 161 5.1.3 Efficiency of sperm separation from seminal plasma and infectious organisms 162 5.2 General principles 162 5.3 Simple washing 163 5.3.1 Reagents 163 5.3.2 Procedure 163 5.4 Direct swim-up 164 5.4.1 Reagents 164 5.4.2 Procedure 164
vi 5.5 Discontinuous density gradients 165 5.5.1 Reagents 165 5.5.2 Procedure 166 5.6 Preparing HIV-infected semen samples 166 5.7 Preparing testicular and epididymal spermatozoa 167 5.7.1 Enzymatic method 167 5.7.2 Mechanical method 167 5.7.3 Processing sperm suspensions for intracytoplasmic sperm injection 167 5.8 Preparing retrograde ejaculation samples 168 5.9 Preparing assisted ejaculation samples 168 Chapter 6 Cryopreservation of spermatozoa 169 6.1 Introduction 169 6.2 Semen cryopreservation protocols 172 6.2.1 Standard procedure 172 6.2.2 Modified freezing protocols for oligozoospermia and surgically retrieved spermatozoa 175 6.2.3 Labelling of straws and records 176 PART III. QUALITY ASSURANCE Chapter 7 Quality assurance and quality control 179 7.1 Controlling for quality in the andrology laboratory 179 7.2 The nature of errors in semen analysis 179 7.3 Minimizing statistical sampling error 180 7.4 The quality assurance programme 182 7.5 Laboratory procedures manual 182 7.6 Internal quality control 182 7.6.1 Purchased QC samples 183 7.6.2 Laboratory-made QC samples 183 7.6.3 Stored samples (purchased or laboratory-made) 183 7.6.4 Fresh QC samples (laboratory-made) 184 7.7 Statistical procedures for analysing and reporting within- and among-technician systematic errors 185 7.7.1 The X bar chart 185 7.7.2 The S chart 188 7.8 QC for percentages 189 7.9 Assessing X bar and S charts 189 7.9.1 How to recognize out-of-control values 189 7.9.2 Causes of out-of-control values 190 7.9.3 Responses to out-of-control values 191 7.10 Statistical procedures for analysing and reporting among-technician variability 191 7.10.1 Comparing results from two or more technicians 191 7.10.2 Monitoring monthly means 194 7.11 External quality control and quality assurance 194 7.11.1 Assessment of EQC results 196 7.11.2 Responses to out-of-control results 197 7.12 Frequency and priority of quality control 197
Page 1 and 2: WHO laboratory manual for the Exami
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42 PART I Semen analysis Table 2.4
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44 PART I Semen analysis The concen
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46 PART I Semen analysis Plac
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48 PART I Semen analysis 2.11 When
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50 PART I Semen analysis Table 2.5
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52 PART I Semen analysis 2.11.2 Ass
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54 PART I Semen analysis Box 2.13 V
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56 PART I Semen analysis the concen
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58 PART I Semen analysis Fig. 2.10
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60 PART I Semen analysis Note 3: Be
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62 PART I Semen analysis Note 1: If
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64 PART I Semen analysis 2.14.2.2 F
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66 PART I Semen analysis 2.14.3.3 S
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68 PART I Semen analysis Spermatozo
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72 PART I Semen analysis 10 microns
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100 PART I Semen analysis Comp
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102 PART I Semen analysis 2.17.6 As
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104 PART I Semen analysis 2. Remove
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106 PART I Semen analysis 2.18.1.6
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108 PART I Semen analysis Nuclear s
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110 PART I Semen analysis 9. Repeat
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112 PART I Semen analysis 1. Mix th
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114 PART I Semen analysis 4. Includ
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116 PART I Semen analysis The SDI
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118 PART I Semen analysis 3.2.2 Rea
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120 PART I Semen analysis 5. Calcul
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122 PART I Semen analysis 3.3 Inter
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124 PART I Semen analysis Box 3.2 V
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126 PART I Semen analysis mid-cycle
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130 PART I Semen analysis Poor-qual
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132 PART I Semen analysis 2. Reject
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134 PART I Semen analysis 4. Replic
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136 PART I Semen analysis 7. Stop i
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138 PART I Semen analysis recording
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140 PART I Semen analysis 3.5.3 Use
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142 CHAPTER 4 Research procedures W
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144 PART I Semen analysis 6. Phorbo
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148 PART I Semen analysis 3. NaCl,
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152 PART I Semen analysis 4.4.2.4 Q
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154 PART I Semen analysis 5. Establ
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156 PART I Semen analysis 4.5.1.7 C
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PART II. Sperm preparation
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162 PART I I Sperm preparation and
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164 PART I I Sperm preparation Note
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166 PART I I Sperm preparation 6. G
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168 PART I I Sperm preparation 5. P
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170 PART I I Sperm preparation Box
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172 PART I I Sperm preparation Note
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179 CHAPTER 7 Quality assurance and
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CHAPTER 7 Quality assurance and qua
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References
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206 References Auger J, Eustache F
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208 References Chohan KR et al. (20
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210 References Davis RO, Katz DF (1
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212 References Grimes DA, Lopez LM
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214 References Kruger TF et al. (19
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216 References Menkveld R, Kruger T
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218 References Rossi AG, Aitken RJ
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220 References WHO (1996). Task For
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223 APPENDIX 1 Reference values and
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APPENDIX 1 Reference values and sem
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227 APPENDIX 2 Equipment and safety
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APPENDIX 2 Equipment and safety 229
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APPENDIX 3 Microscopy 235 Note: If
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APPENDIX 3 Microscopy 237 Note: If
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APPENDIX 4 Stock solutions 239 Note
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APPENDIX 4 Stock solutions 241 A4.8
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APPENDIX 4 Stock solutions 243 Note
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245 APPENDIX 5 Cervical mucus A5.1
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APPENDIX 5 Cervical mucus 247 When
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APPENDIX 5 Cervical mucus 249 A5.3.
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251 APPENDIX 6 Record forms for sem
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APPENDIX 6 Record forms for semen a
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APPENDIX 7 Sampling errors and qual
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271 APPENDIX 8 National external qu
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