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--- protocols:cryo [2019/11/01 10:52]
rubin created
+++ protocols:cryo [2023/04/27 13:41] (current)
ron
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 ====== Cryopreservation and In Vitro Fertilization (IVF) Protocols ======
-===== Current Protocols =====
+  *  {{ :protocols:zirc_e400_rmmb_sperm_cryopreservation_ivf_protocol_04.24.2023.pdf |ZIRC E400/RMMB Sperm Cryopreservation & IVF Protocol}}
-  * ZIRC E400/RMMB Sperm Cryopreservation & IVF Protocol by Matthews et al. V.1.2_2017_07_18 {{:protocols:cryo:zirc_rmmb_freezing_protocol.pdf|PDF}}
+      * Complete description of ZIRC's current cryopreservation & IVF protocols
-    * A step-by-step description of ZIRC's current cryopreservation & IVF protocols
-  * Calibration Curve and Sperm Density Calculator {{:protocols:cryo:zirc_nanodrop_sperm_density_calc.xls|XLS}}
+  * [[:protocols:cryo:step-by-step_cryopreservation_protocol]]
-    * Sperm Density Calculator for use with Nanodrop Spectrophotometer concentration determination
+      * Abridged version of ZIRC's cryopreservation & IVF protocols
-===== Previous Protocols =====
+  *  {{ :protocols:zirc_nanodrop_2000_calibration_curve_and_sperm_density_calculator.xlsx | ZIRC NanoDrop 2000 Calibration Curve and Sperm Density Calculator}}
-  * Cryopreservation of zebra fish spermatozoa using methanol by Harvey et al. 1982 {{:protocols:cryo:v.60-zoology-pp1867-70.pdf|PDF}}
+      * ZIRC's NanoDrop 2000 hemocytometer-generated standard curve and sperm concentration calculator for use with OD400 measurements for stripped sperm in E400 extender
-    * [[http://pubs.nrc-cnrc.gc.ca/cgi-bin/rp/rp2_desc_e?cjz|The Canadian Journal of Zoology]] has kindly permitted us to provide a downloadable document of the original article by Harvey et al., (1982) that describes cryopreservation for zebrafish. All copyrights remain with the Canadian Journal of Zoology. You can download the article for your own use. Please [[http://pubs.nrc-cnrc.gc.ca/eng/policy/index.html#copyright|inform yourself here]] about copyright issues.
-  * Sperm Cryopreservation by Walker and Streisinger
+  *  {{ :protocols:diluting_pooled_sperm_based_on_nanodrop_cell_counts_04.24.2023.pdf |Diluting Pooled Sperm based on Nano Drop Cell Counts}}
-    * The laboratory protocol that was adapted from Harvey et al. by Walker and Streisinger can be found in [[http://zfin.org/zf_info/zfbook/chapt7/7.6.html|the Zebrafish Book, Chapter 7]].
+      * Detailed description of how to use a NanoDrop 2000 spectrophotometer for measuring sperm cell concentrations and how to use the Excel Pooled Sperm Dilution Worksheet
-  * Sperm Cryopreservation by Draper and Moens. 2004
+  * {{ :protocols:pooled_sperm_dilution_worksheet_blank.xlsx |Pooled Sperm Dilution Worksheet}}
-    * View Draper and Moens' [[http://authors.fhcrc.org/323/|complete article and protocol video]] (A high-throughput method for zebrafish sperm cryopreservation and in vitro fertilization) on the Fred Hutchinson Cancer Research Center web site. This is the protocol on which we base our procedures  here at ZIRC (see 4, below).
+      * A blank Excel sperm dilution worksheet for use with Nanodrop 2000 cell counts as described in Diluting Pooled Sperm based on NanoDrop Cell Counts
-  * Cryopreservation and In Vitro Fertilization at the ZIRC by Carmichael et al. 2009 {{:protocols:cryo:zirccryo_ivf.pdf|PDF}}
-    * This review provides a step-by-step description of the Draper and Moens method (see 3, above) as it is currently applied at the ZIRC. In addition, we include detailed notes with background, advice, and additional information. The authors, and ZIRC in general, wish to express their gratitude to Cecilia and Bruce for teaching us the protocol and for their continued advice and support.

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