MOPS and HEPES can be combined as a gamete buffer

MOPS, 3-(N-Morpholino)propanesulfonic acid, is an important biological buffer, the buffer range is 6.5~7.9; HEPES, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, is also a hydrogen ion buffer which has a Good buffering capacity in the pH range of 6.8~8.2 and can control a constant pH for a long time. They are both widely used in biological and chemical experiments. When the two combine to act as a gamete buffer, what are the advantages?

The importance of gamete buffer

Gametes refer to mature cells produced by the reproductive system when the organism is sexually reproducing, referred to as germ cells. Gametes are divided into male gamete and female gamete. The female gametes of animals and plants are usually called egg cells (ova, or egg), and male gametes are called sperm. The gamete buffer is a buffer containing antibiotics, basic ions and various energetic substances. It is used for washing eggs and sperm outside the incubator to prepare for IVF (in vitro fertilization). Drastic fluctuations in the environment can severely impair the viability of gametes, which can have a devastating effect on the quality and viability of the embryo. The success of subsequent operations in the IVF depends on whether the homeostasis of the oocyte can be maintained. Gamete buffers maintain the metabolism of oocytes and reduce the effects of stress on gametes, thus preparing for fertilization.

Traditional gamete buffer
Currently, MOPS, HEPES, or sodium bicarbonate-carbon dioxide buffer pairs are often used in traditional gamete buffers. Although MOPS and HEPES are widely used in the field of assisted reproduction, the effects of temperature on the pKa values of both are often ignored, as well as the effects of MOPS on illumination. Therefore, the potential hazard of buffering on gamete and embryo quality prompted researchers to reassess the pH buffer system currently used for clinical in vitro fertilization.

Therefore, a more stable pH-matching buffer system was developed, as well as a gamete buffer prepared using the system. The gamete buffer of the present invention reduces the effect of pH on the activity of gametes, and the gametes washed by the gamete buffer have a greatly improved blastocyst formation rate and uniform morphology in the late embryo test.

New gamete buffer

Both MOPS and HEPES are zwitterions based on aminosulfonic acids, which act as ampholytes and provide a relatively stable pH gradient. The inventors have found that the combination of MOPS and HEPES can control the pH in a narrower range and have a stronger buffering capacity than a single buffer pair at the same concentration. Therefore, using the MOPS and HEPES buffer system, the stress effect of gametes can be reduced and prepared for subsequent IVF. At the same time, the same concentration of MOPS and HEPES buffer system reduces the cytotoxicity of the buffer pair compared with the single buffer pair, and is more conducive to the viability of the gametes.

The specific plan is as follows:

The solvent is water, including MOPS and HEPES buffer systems; it also contains sodium, potassium, calcium, magnesium ions, as well as glucose, sodium pyruvate, albumin and non-essential amino acids. In the preferred solution:

• The above gamete buffer components are: 90.24~110.18mmol/L sodium chloride, 0.43~0.52mmol/L magnesium chloride, 4.267~5.013mmol/L potassium chloride, 0.1522~0.2567mmol/L magnesium sulfate, 0.13~0.30mmol/ L-calcium pantothenate, 0.3513~0.3958 mmol/L potassium phosphate, 21.0~21.6 mmol/L calcium lactate, 8~12 mmol/L HEPES, 8~12 mmol/L MOPS, 3.85~4.30 mmol/L sodium bicarbonate, 0.3129~0.3501 mmol/L-sodium pyruvate, 2.5~3 mmol/L glucose, 0.95~1.06 mmol/L alanyl glutamine, 0.09~0.10 mmol/L L-taurine, 5~10 ug/mL gentamicin sulfate, 5 mg/mL human serum albumin and 0.5~1.5mmol/L human non-essential amino acids (glycine, L-alanine, L-valine, L-serine, L-asparagine, L-aspartate , L-glutamic acid).
• The pH of the gamete buffer is 7.25~7.35.
• The osmotic pressure of the gametophysic buffer is 285~295 mOsm/kg.
• The endotoxin content of the gamete buffer is <0.25 EU/mL.

Compared with the conventional gamete buffer liquid, the endotoxin content of the buffer is lower, which reduces the toxicity to the gametes during use; the pH range is narrower and more stable, and the gametes can better maintain the steady state and reduce the vigor activity of the gametes when used. The embryo morphology is also satiation, which is conducive to embryo formation and development, thus preparing for in vitro fertilization.

Edited by Suzhou Yacoo Science Co., Ltd.