Introduction

The first tissue culture media were developed nearly 150 years ago by Ludwig and Ringer. These were simple salt solutions, which were initially based on the chemical properties of blood serum. Knowledge was gained during those early years about the biochemistry of metabolism in mammals and humans, particularly osmolarity, pH and temperature. This knowledge provided the basis for making many changes to the salt solutions that were already in use. The second generation of culture media was...

Notes

Incubators for maintaining the atmosphere with reduced oxygen are often named triple-gas incubators, but this term is a misnomer since 5 CO2 in the air is also triple gas (CO2, O2, and N2). Triple-gas incubators have two gas sensors. To ensure more stable conditions, an infra red (IR) sensor is recommended, and not a thermal conductivity (TC) CO2 sensor. The IR sensor works independently of humidity and temperature. The oxygen sensor is more sensitive and, in comparison with the CO2 sensor, its...

Materials

Tissue culture dishes (35 x 10 mm). 2. Tissue culture dishes Falcon 3002 (60 x 15 mm). 5. Centrifuge conical graduated tubes. 7. B2 medium (with calf serum). 9. Hanks balanced solution w o Ca and Mg. 10. Vero cells. The cells were either obtained from ATCC or WHO collection (UK), at sub-passage 120-134, in a frozen tube containing 25 million cells. A certificate of quality including all the tests for viruses and electron microscopy is given. 13. Chlamydia (BioMerieux Kit 5532 1). 14. Mycoplasma...

Table 3 continued

'Cleavage indicates embryos after fertilization to the time of compaction. 8c denotes 8-cell embryos usually around the time of compaction, and Bl denotes blastocyst. Media listed as cleavage and 8c to Bl are paired as sequential media. Media listed as all stages are single media that can be used for culture from fertilized egg to blastocyst bOsmolarity ranges for media from these companies were obtained from the Web sites listed in Table 1 for each individual medium cPatrick Quinn, PhD, HCLD,...

Acknowledgments

The author would like to thank Kimball Pomeroy for sharing his insights into and experiences with mammalian embryo culture, and his critical reading of this manuscript. The author would also like to acknowledge Biogenics (Napa, CA) and John Haynes (SPI, Inc Canton, TX) for samples of, and information pertaining to the Conical Embryo Culture Plate, and Hong Wei (Trans Ova Genetics, Sioux Center, IA) for sharing unpublished data re the Conical Embryo Culture Plate Hamilton Thorne Inc. (Beverly,...

Embryo Culture Notes

For accuracy, care must be taken in setting up of patient dishes. To accurately determine the uptake of energy sources such as pyruvate, glucose or amino acid from spent culture media, a degree of care must be taken when setting up culture dishes as even small variations in drop size between embryos will result in significant changes to nutrient levels within the drop after culture. Therefore, all culture dishes should be prepared utilizing an accurate pipette (such as a Gilson multi-pipette)...

Methods

How to Establish The transition from high- to low-oxygen culture can be a challenging Low-Oxygen Culture project, both in financial terms and in the organization of work in System the laboratory. The major problem occurs in laboratories which are already equipped with a large number of incubators that are not able to regulate oxygen. A frequently asked question in this case is whether it is possible to introduce a low-oxygen culture with existing incubators. In this case, the use of the...

Expression of growth factors within the reproductive tract in relation to the menstrual cycle

Various cell types and fluids within the reproductive tract Growth factors Phase of the menstrual cycle Endometrium Leukemia inhibitory factor (LIF) Activin A by the epithelial cells (92) Colony stimulating factor (CSF) (87) Insulin-like growth factor 1 primarily Insulin-like growth factor 2 (146, 147) Platelet derived growth factor (PDGF) Various cell types and fluids within the reproductive tract FGF and fibronectin (83) Insulin like growth factor-1 (73) Insulin like growth factor-2 (73)...

Table 2 continued

Flush collection media, heparin, searching dishes, warm surfaces (stages, laminar flow hoods), time, technical skill Oocyte washing medium, warm surfaces (stages laminar flow hoods), exposure to light (microscopes), pipettes, culture dishes, incubator performance (temperature, gas phase, recovery), oocyte culture medium, oocyte culture oil, hyaluronidase, stripping pipettes, technical skill Cleavage-stage culture medium, embryo culture dishes, culture oil, incubator performance (temperature,...

Materials Used In Embryo Culture

Appropriate embryo tested culture dishes. 3. Fertilization, cleavage, and blastocyst media. Comparison of sequential media and single-step medium culture protocols Factors Sequential media Single-step medium Comparison of sequential media and single-step medium culture protocols Require change of medium around D3 of culture Can be used for complete preimplanta-tion period with no medium change Blastocyst medium does not contain EDTA that may reduce glycolysis Contains EDTA that may compromise...

Gynemed 501 Culture Media How Widely Used

The human oocyte fertilizes in the ampullary region of the fallopian tube in vivo, and the newly formed zygote migrates through the isthmus, traverses the uterotubal junction around day 4 at the morula stage, and subsequently implants in the uterus around day 6, at the stage of blastocyst (1-3). Analyses of energy substrate concentrations in fluid collected from nonpregnant women at various stages of the menstrual cycle have shown that these concentrations differ between the fallopian tube and...

Voc Packaging Embryo

The importance of air quality in the culture of human gametes and embryos began to emerge in the 1990s when reproductive biologists performing human in vitro fertilization IVF procedures began to associate episodic declines in clinical outcome with construction or renovation in or near the laboratory. Early anecdotal information led to the theory that laboratory air could be contaminated with volatile compounds or particulates derived from construction activity, outgassing of new materials, or...

Gynemed Gm501 Composition

Salts in Human The overwhelming majority of the solutes present in embryo Embryo Culture Media culture media consists of inorganic ions from salts. All commercial human embryo culture media share the same six inorganic ions Na , K , Cl-, Ca2 , Mg2 , and SO42-, which are the same ones that were included in the first defined mouse embryo culture media formulated in the 1950s 1, 2 . Most media also contain PO42-, although several that were created specifically for eggs and cleavage stage...

Does Vitrolife Ovoil Absorbs

Mineral oil has been a standard component of embryo culture since the early 1960s. Following the initial success of culturing cleavage stage embryos to blastocyst in test tubes 1, 2 , two students in the laboratory of John Biggers at the University of Pennsylvania, Ralph Gwatkin 3 and Ralph Brinster 4 , developed a method which allowed more frequent observation of embryos that entailed the use of mineral oil placed over the top of media in petri dishes 5 . Though culturing in test tubes is...

Power and sample size estimates to detect birth rate differences among four quality grade groups df3

Effect size e Alpha a Beta Power 1 - Chi-square c2 Sample size n Power and sample size estimates to detect birth rate differences among seven quality grade groups df 6 Effect size e Alpha a Beta Power 1 - Chi-square c2 Sample size n Power and sample size estimates to detect birth rate differences among seven quality grade groups df 6 Clearly, the experimental isolation of culture medium formulation as the sole variable contributing to altered outcome following IVF is extremely difficult and,...

Introduction Of Embryo Culture

Current IVF laboratory practice is to supplement all commercial IVF embryo culture media with some protein. Actually, most if not all mainstream commercial media manufacturers will provide Gary D. Smith et al. eds. , Embryo Culture Methods and Protocols, Methods in Molecular Biology, vol. 912, DOI 10.1007 978-1-61779-971-6_8, Springer Science Business Media, LLC 2012 complete or ready to use protein-supplemented products. It may be prudent to consider the history and origins of culture media...