CN104480063A - Production method of ICSI operation vessel - Google Patents

Abstract

The invention discloses a production method of ICSI operation vessel. The production method comprises the following steps under the following conditions: sucking PVP solution by a suction tube, forming two 10 microliter circular micro-droplets above the ICSI vessel and forming a strip-shaped PVP micro-droplet with a length of 2.5cm and a width of 0.5cm under the ICSI vessel; sucking about 0.1ml of a culture medium containing sperms, and forming a strip-shaped micro-droplet with a length of about 2.0cm and a width of about 0.5cm at one side of the strip-shaped PVP micro-droplet; sucking a buffer culture medium, and forming three to five 10 microliter circular micro-droplets containing an ovum at the other side of the strip-shaped PVP micro-droplet; sucking the buffer culture medium, and forming one 10 microliter circular micro-droplet under the strip-shaped PVP micro-droplet; sucking one sperm from the strip-shaped micro-droplet of the culture medium containing sperms by a microscopic puncture needle, placing the sperm in the strip-shaped PVP micro-droplet, and immobilizing the sperm; fixing the ovum by a microscopic fixation needle so that the polar bodies of the ovum can be located at a six-o'clock position or a twelve-o'clock position; injecting the sperm in the ovum by the microscopic puncture needle with the sperm from a three-o'clock position.

Description

A kind of ICSI operates ware making method

Technical field

The present invention relates to and a kind ofly can look at short notice and draw the method for sperm, be specifically related to the making method that a kind of ICSI operates ware.

Background technology

In monosperm ooecium slurry, injection technique (ICSI) makes countless severe oligospermia, Obstructive azoospermia patient has oneself offspring, ICSI operating process must operate on ware at ICSI and complete, traditional IC SI operates ware by ICSI ware, the circular droplet of PVP, PVP bar shaped droplet and buffer culture medium droplet composition, process is drawn by the substratum containing sperm after 1.0 μ about l to be added in PVP bar shaped droplet braking, complete ICSI operation, but the situation that after running into some washings, sperm concentration is few, this method is used to be difficult to find sperm at short notice, the ICSI operating time is extended greatly, ovum is exposed to incubator outside for a long time, constant temp can not be kept, humidity and CO2 concentration, the ultrastructure of ovum can be damaged unavoidably, affect its fertilization and potentiality of development, if the amount adding substratum is excessive, then sperm sum increases, relatively easily find sperm, but sperm can be caused in PVP solution to be in levitated state, puncture needle is difficult to capture and braking sperm, the ICSI operating time can be extended equally, and some the sperm impurity in substratum can pollute PVP solution.

Some IVF laboratories suggestion severe oligozoospermia and obstructive without sperm patient repeatedly get essence afterwards freezen protective to increase sperm total amount, this method drawback is: repeatedly cryopreservation sperm can increase treatment cost, its freezing rear resuscitation effect of sperm that current density is very low is undesirable, and whether refrigerating fulid has liquid nitrogen in damage, refrigerating process whether can produce pollution to sperm, use its security of sperm of recovery how at present still without final conclusion to sperm.

Summary of the invention

The object of the invention is the technical problem operating ware existence in order to solve existing ICSI, the invention provides a kind of new ICSI and operating ware making method.

Principle of the present invention is: the sperm mediun bar shaped droplet that present method is done is about 0.1ml, large owing to measuring, sperm in substratum adds up to about 100 times of traditional method, its traditional method amount is 1.0 μ about l, the measured sperm of substratum endoplasm can move to bar shaped droplet edge, because sperm total amount increases, find and capture the sperm time greatly to shorten.

A kind of ICSI provided by the invention operates ware making method, step and condition as follows:

Step one: draw PVP solution with suction pipe, does two circular droplets of 10 μ l above ICSI ware;

Step 2: do the bar shaped PVP droplet that length and width are respectively 2.5 cm, 0.5cm immediately below droplet;

Step 3: draw containing the substratum 0.09-0.2ml of Sperm pellets, doing long in PVP bar shaped droplet side is 1.8-2.2cm, and wide is the bar shaped droplet of 0.4-0.6cm;

Step 4: draw buffer culture medium and do 3-5 the circular droplet of 10 μ l at PVP bar shaped droplet opposite side, put into ovum in droplet;

Step 5: draw buffer culture medium and do 1 circular droplet of 10 μ l immediately below PVP bar shaped droplet, namely wash pin;

Step 6: draw 1 sperm with microscopical puncture pin in the substratum bar shaped droplet containing sperm, put into PVP bar shaped droplet, braking sperm;

Step 7: fix ovum with microscope fixing pin, makes its polar body be in or 12 positions at 6;

Step 8: with the microscopical puncture pin of sperm from 3 positions by sperm injection in ovum, namely complete ICSI and operate ware and make.

The beneficial effect of employing technique scheme is:

Injection operation ware making method in a kind of new monosperm ooecium slurry provided by the invention, is a kind of simple, practical making method, is applicable to severe oligospermia, epididymis gets essence and testis gets the ICSI operation of the special seminal fluid such as essence.The good sperm of absorption quality also can be found to complete ICSI operation in the short period of time, reduce ovum and cruelly leak the time outward at incubator.Reduce the change of temperature, humidity and CO2 concentration to greatest extent to Ultrastructural damages such as ovum spindle bodys.

The present invention utilizes each sperm to greatest extent, is particularly suitable for the situation that sperm quantity is few.

Method of the present invention is applicable to severe oligospermia, epididymis gets essence and testis gets the operations of ICSI such as essence, greatly shortens sperm and captures and braking time.

Accompanying drawing explanation

Fig. 1 is the ICSI operation ware schematic diagram that ICSI of the present invention operates ware making method.

In figure: 1-ICSI ware, 2-buffer culture medium circle drips, the bar shaped of 3-PVP solution is dripped, 4-Sperm pellets bar shaped droplet, 5-PVP solution circle drips.

Embodiment

Below the present invention is described in further detail:

A kind of ICSI operates ware making method, step and condition as follows:

Step one: draw PVP solution with suction pipe, does two circular droplets of 10 μ l above ICSI ware;

Step 2: do the bar shaped PVP droplet that length and width are respectively 2.5 cm, 0.5cm immediately below droplet;

Step 3: draw containing the substratum 0.09-0.2ml of sperm sperm, doing long in PVP bar shaped droplet side is 1.8-2.2cm, and wide is the bar shaped droplet of 0.4-0.6cm; All Sperm pellets are all in strip droplet 0.09-0.2ml, so just do not waste each sperm;

Step 4: draw buffer culture medium and do 3-5 the circular droplet of 10 μ l at PVP bar shaped droplet opposite side, put into ovum in droplet;

Step 5: draw buffer culture medium and do 1 circular droplet of 10 μ l immediately below PVP bar shaped droplet, namely wash pin;

Step 6: draw 1 sperm with microscopical puncture pin in the substratum bar shaped droplet containing sperm, put into PVP bar shaped droplet, braking sperm;

Step 7: fix ovum with microscope fixing pin, makes its polar body be in or 12 positions at 6;

Step 8: with the microscopical puncture pin of sperm from 3 positions by sperm injection in ovum, namely complete ICSI and operate ware and make.

ICSI ware of the present invention is produced by BD Falcon company of the U.S., article No.: 353001.

Operation of the present invention is dripped and is used U.S. Quinn ' s to cushion nutrient solution, article No.: ART-1023.

Substratum containing sperm of the present invention is exactly the Sperm pellets after washing.

7%PVP solution of the present invention is that SAGE Media company of the U.S. produces, article No.: ART-4005-A.

Microscopical puncture pin of the present invention is that PIPETTE company of the U.S. produces, article No.: S0206R.

Microscope fixing pin of the present invention is that PIPETTE company of the U.S. produces, article No.: S0310Q.

Embodiment

Between in September ,-2014 in May, 2013, get to serious few essence, epididymis 316 pieces of mature egg sub-line ICSI that essence and testis get 28 its spouses of patient of essence altogether to operate, traditional IC SI operation ware is used to carry out 173 pieces, the ovum of ICSI operation, average operation time is 217 ± 7.0 seconds, new ICSI is used to operate 143 pieces, ware ovum, average operation time 122 ± 9.0 seconds, between two groups, there were significant differences, and all operations is same technician and completes.

Claims (1)

1. ICSI operates a ware making method, it is characterized in that: step and condition as follows:

Step one: draw PVP solution with suction pipe, does two circular droplets of 10 μ l above ICSI ware;

Step 2: do the bar shaped PVP droplet that length and width are respectively 2.5 cm, 0.5cm immediately below droplet;

Step 3: draw containing the substratum 0.09-0.2ml of sperm sperm, doing long in PVP bar shaped droplet side is 1.8-2.2cm, and wide is the bar shaped droplet of 0.4-0.6cm;

Step 4: draw buffer culture medium and do 3-5 the circular droplet of 10 μ l at PVP bar shaped droplet opposite side, put into ovum in droplet;

Step 5: draw buffer culture medium and do 1 circular droplet of 10 μ l immediately below PVP bar shaped droplet, namely wash pin;

Step 6: draw 1 sperm with microscopical puncture pin in the substratum bar shaped droplet containing sperm, put into PVP bar shaped droplet, braking sperm;

Step 7: fix ovum with microscope fixing pin, makes its polar body be in or 12 positions at 6;

Step 8: with the microscopical puncture pin of sperm from 3 positions by sperm injection in ovum, namely complete ICSI and operate ware and make.