Ke-Hui Cui M.D., Ph.D.

Savannah, Georgia, 31405, U.S.A.

March 15th, 2019

Email: khcui72@hereditics.net

* This historical document was one of two attached papers sent to CDC, FDA and ASRM on March 15th, 2019. Another attached paper was "Origin of life, heredity, differentiation and human reproduction: control of overpopulation and prohibition of subhuman reproduction" (Shortened as "Origin of life"), in which Hereditics was first time set up. Immediately US Congress positively recognized the new scientific theory - Hereditics. According to these two important papers, US Congress issued "Personhood Concern" to ASRM about the severe issue of "preimplantation genetic testing for aneuploidy" in March, 2019. Without cognition of new theory in Hereditics, Cytohetics and Epicytohetics (rather than Genetics), it is not possible for physicians to understand why PGT-A is neither scientific nor safe, for geneticists to explain why mosaic embryos may produce many normal karyotyping children, and for embryologists to understand why the presence of aneuploidy is a normal condition in human embryology. Thus, study of hereditics are urgently needed. Due to this reason, this document "Preimplantation genetic test for aneuploidy is lack of scientific bases and is not safe" was rewritten into two detailed educational papers to specially explain Cytohetics and Epicytohetics individually: "Cytohetics: preimplantation genetic testing for aneuploidy is not scientific" and "Epicytohetics: preimplantation genetic testing for aneuploidy is not safe". Another attached paper "Origin of life" was also rewritten into an educational paper "Hereditics and origin of life". All of these papers have been published at https://www.hereditics.net. Other important papers and documents in the field of life and health science, including "Ban subhuman reproduction", "Control of overpopulation", "MRT syndrome", "Legal discussion related to children born with embryo biopsy syndrome and laser radiation syndrome",  etc. have been or will be published on the hereditics.net.

Abstract

Presence of aneuploidy cells in human embryonic stages is an important and positive phenomenon of embryonic physiology for differentiation and for faster implantation. Preimplantation genetic test for aneuploidy (PGT-A) or preimplantation genetic screening (PGS) is to screen embryos to find out aneuploidy cells (or abnormal DNA). It is not following parents’ history. Clinical results from two papers showed: 26 embryos containing aneuploidy cells were transferred and they developed to 11 normal karyotyping babies. The heredity ability of those diagnosed aneuploidy cells (i.e. DNA) was not persistent in these live babies. Genetics about DNA to be the unique hereditary materials could not be used to explain this result. However, when following parent’s genetic history, preimplantation genetic diagnosis (PGD) was performed. In 3755 pregnancy, only 34 misdiagnosis cases (0.9%) were reported. The results showed the embryos were keeping on their normal diagnosed heredity (>99.0% correct rate). Whether following patient’s history or not, the PGT results were significantly different (p<0.0001) in prediction of the heredity. If not following patient’s history, the diagnosed DNA (aneuploidy cells) will not show its heredity characteristic. Evolution history showed: origin of heredity was not DNA, but a combination of RNA and lipid membrane. Patient’s history is closely related to how perfect the hereditary control system is. Hereditary diseases are the combination results of both defects of DNA and heredity control system. To study the heredity function of cytoplasm is imperative. To use aneuploidy-producing techniques (embryo biopsy, laser radiation and freezing) to prevent aneuploidy is not scientific and not safe in PGT-A.

History of PGS

After failure in polymerase chain reaction (PCR) techniques with misdiagnosis of a female embryo, a boy with a hereditary disease was born in 1991. Dr. Handyside introduced fluorescent in-situ hybridization (FISH) techniques in sex determination for prevention of X-linked hereditary diseases. “Aneuploidy” phenomenon in human embryos was immediately emerged with FISH techniques. “Aneuploidy” debate was initiated. It was claimed by Dr. Verlinsky that “60% of human embryos were aneuploidy”, and preimplantation genetic screening (PGS) started. The hypothesis of PGS was that if all embryos obtained by IVF were screened for aneuploidy prior to transfer, implantation and pregnancy rates would improve and miscarriage rates decrease (Munné et al., 1993). FISH techniques are full of error to produce artifact of “aneuploidy”, in which only one side of the Chromosomes sticking of the glass will produce many artifact. Later, array comparative genomic hybridization (aCGH) and next generation sequencing (NGS) emerged in place of FISH techniques.

Aneuploidy embryo means that all of the cells in that embryo are aneuploidy. People are not possible using PGT to diagnose all of the cells of an embryo and transfer an empty zona to produce a life. People can only biopsy 1/8 of cells in 8-cell stage or up to 5 cells in blastocyst for PGT to represent the whole embryo. Thus their results can not confirm whether the said embryo is aneuploidy or not, but only confirm whether there is/are any “aneuploidy” cell to be found. After trophectoderm biopsy, people began to say that “60% of human embryos were mosaicism” rather than “aneuploidy”.

In 2015 ASRM meeting, Reprogenetics (recent Cooper Genomics) recommended to use blastocyst biopsy (trophectoderm biopsy) for PGS and elective Single Embryos Transfer (eSET). The eSET is to produce singleton, which is better for mother’s and child’s health, and is warmly welcome by the society. However, it is not the reason to perform the very draumatic techniques including laser radiation, embryo biopsy and freezing for every embryo, of which the majority are healthy embryos. The combination of PGS and eSET was misleading. In 2016, a “Great Leap” of PGS in U.S. happened. The results of massive movement was foreseeable: In 2017 ASRM meeting, “mosaicism tornado” occurred. News reported in New York that two normal babies were born from “mosaicism” embryos, because the mothers insisted transfer of “mosaicism” embryos while no “normal embryo” existed. Why could the “mosaicism” embryos produce the babies with normal rather than mosaic karyotyping? All people’s belief changed when more similar news came out – Transfer of 26 “mosaicism” embryos produced another 11 babies with normal karyotyping (42.3% birth rate) with only 3 miscarrage (Greco, et al., 2015, Gleicher, et al. 2016). The fact that 11 normal babies born from “abnormal mosacism” embryos confirmed that all of the theory and practice of PGS failure. The majority of diagnosed aneuploidy cells with abnormal DNA structures do not contain characteristics of heredity in the born babies. Diagnosis of PGS was not functioning anymore. Why does the diagnosed DNA (of the aneuploidy cells) from biopsy not keep its heredity? The credibility of PGS were all vanished (Gleicher, et al., 2018), from its most basic points: when to biopsy, how to biopsy, what instrument to be used for biopsy, how to treat the biopsied cell(s), to high technique question: how to solve allete drop out problem, how to design primers, how to select techniques for diagnoses, etc., to society and ethics questions: how to lecture patients scientifically, how to publish a scientific paper, how to organize our scientific meeting without dictatorship, etc. On the whole, it is the question of “how to deal with corruption” in human reproduction.

The clinical results in U.S. in 2016 got a great setback due to the great leap of PGS. According to Society for Assisted Reproductive Technology (SART) data, the cumulative outcome in U.S. per egg retrieval cycle, in lesser the 35 years old group, Live Births in 2015 was 53.9%, while in 2016 was 47.6%. It was 6.3% drop down, with singletons rate drop down 3.0% (from 42.4% to 39.4%). If without about half of the IVF cycles not involving PGS, the drop down of birth rate would be double. All of the fairy tale (“if all embryos obtained by IVF were screened for aneuploidy prior to transfer, implantation and pregnancy rates would improve and miscarriage rates decrease”) waned when facing the ironic fact. All of the past plenty of “scientific” papers of superiority of PGS with their “data” showed their ugly faces in the authenticity of the greatest scale of clinical malpractice. This serious setback has never been seen in the history of in vitro fertilization. 

Part I: PGT-A (or PGS) is not scientific and it is aimless in heredity

1. Aneuploidy cells exist in our bodies, in fetus and in embryos. Our bodies have plenty of cells dying every day and every hour, in which most of the dying cells are aneuploidy. Should we biopsy ourselves every day everywhere, and freeze the biopsied tissue to transfer later, as now a lot of laboratories biopsy all embryos, freezing before they are transferred to the uterus? Ridiculous. The temporary existence of aneuploidy cells are the physiological mosaic phenomena in our bodies but is not pathological mosaicism. Why are we not mosaicism or not called mosaicism? New scientific research results in recent 10 years showed new heredity control theory: every cell, (it does not matter where it is located, in our bodies, in fetus or in the early embryos,) contains heredity control (security) system, called check points and licensing system (see attached paper “Origin of Life” page 11-13). The check points and licensing system will not allow the aneuploidy cells to cleave and to grow further. When check points and licensing system is in normal condition, the aneuploidy cells will not pass mitotic exit to split, and to be inherited to progeny cells. The aneuploidy cells will be temporarily to be apoptosis and no mosaicism exist. The normal cells will grow and grow. The embryos grow to be fetus and to be our human beings without mosaicism. That is not self-correction, because aneuploidy cells are not possible to restore to be normal cells again. Thus, we should make the different concepts of “mosaic phenomena” and “mosaicism” to be clearly cut at the level of embryology. We should not define an embryo to be mosaicism just depending on the laboratories results of “aneuploidy cell(s) found” or how many percent of aneuploidy cells to be found. Mosaicism is a persistent abnormal growing pathological entity, which hereditary control system is abnormal. But mosaic phenomena are the temporary phenomena, which heredity control system is normal: aging and sick cells will die and new aging and new sick cells will come out, which will not grow further. Only the normal cells will grow into the growing entity because the hereditary control system (check points and licensing system) is normal.

2. Trophectoderm biopsy is not scientific in PGT-A. The embryo at 8 cell stage is topipotent. When embryo grows to blastocyst stage, the cells have differentiated and greatly polarized. In inner cell mass (ICM), the genes in all of the individual cells got about 90% genetic agreement. However, in trophectoderm cells, the genes in all of the individual cells only got about 50% agreement (2018 ASRM paper report). This great difference in genes of individual cells in different tissues was also confirmed by human brain cells. (Different brain cells contain different gene composition). This kind of difference between ICM and trophectoderm is the results of differentiation. The differentiation also changes the check points and licensing system to be great difference in ICM and in trophectoderm. The ICM, which will develop to germ track later, will maintain strict licensing of transcription and translation (Haig. 2016). However, the trophectoderm cells, which will develop to chorion, will produce new and different loose licensing component, (such as an important subunit of the origin recognition complex (Orc 2) is frequently absent and also Cdt1 is restricted) (Lin, et al. 2009). The new loose check points and licensing system allow the trophectoderm cells to differentiate towards syncytium (with multiple nuclei) and chorionic villi direction (Biancotti and Benvenisty, 2011). It is obvious that syncytium cells are not diploid. Thus more aneuploidy cells are found in trophectoderm is not a pathological problem, but is the normal functions and results from the new and loose check points and licensing system (Hong and Stambrook, 2004) in trophectoderm differentiation. Experimental results showed: 8 embryos were biopsied on day 3 and were confirmed as normal 2N. When they were grown to blastocysts and were analyzed in individual cells, all of them could not showed 100% normal 2N, in which there were three blastocysts showed tetraploid (4N) syncytium cell results. In that experiment, in total 21 blastocysts, there were nine blastocysts containing 4N chromosomes (42.8%) to show the differentiation of villi direction (Baart, et al., 2004; Baart, et al., 2006) It is a positive phenomenon while some aneuploidy cells contain more invasive function for villi to implant into mother’s endometrium (Gleicher, et al. 2016), because “aneuploidy-independent modulation of the microtubule cytoskeleton enhance directional migration and invasion” (Ogden, et al., 2013). Only the villi tissue develops fast enough in cell number, with abnormal nuclei and abnormal aneuploidy cells, so that villi can finish its hard task: invade into the mother’s endometrium faster to obtain enough nutrition to support the fast growing embryo and fetus, and to prevent ICM and embryonic soma from abnormal functioning due to lacking of nutrition. Gynecological data showed: under malnutrition condition, the loose check points and licensing system will lead to real pathological results: hydatidiform mole or choriocarcinoma, which are the aneuploidy entity. When tested 42 human blastocysts by individual cells, almost all of the blastocysts but one (98%) contained some aneuploidy cells (Daphnis, et al., 2005). It confirmed that embryos containing some aneuploidy cells are physiological phenomenon in blastocyst stage. It is nothing related to fetal mosaicism except very low (0.2%) opportunity. Those aneuploidy cells and syncytial cells in the villi will not result in fetal mosaicism and will be gone with the placenta after the baby is born. Those aneuploidy cells and syncytium cells from trophectoderm biopsy are nothing to do with the ICM karyotyping, because the check points and licensing system in trophectoderm cells and the system in ICM cells are greatly different after differentiation. Encourage using trophectoderm biopsy on day 5 in place biopsy on day 3 to find more aneuploidy cells and claiming them as fetal mosaicism and then discarding them, is a kind of business idea to take advantage of the patients. It is lack of the basic scientific knowledge. Trophectoderm biopsy is in the wrong time to perform biopsy at the wrong location with wrong aim and to give wrong diagnoses and wrong treatment. That is the strong reason for the government to step in and ban the clinical malpractice.

3. One paper reported: In 72472 pregnancies, chromosomal mosaicism was 2.1% in chorionic villi samples, while 0.084% in amniotic fluid with total 0.2% aneuploidy. That is: mosaicism in the fetus is lesser than 0.1% (Grati, et al. 2018). Mosaicism in babies is rare are the common sense in all of the Ob/Gyn doctors in the world for 7 decades. It is not safe and not scientific (for prevention of 0.2% aneuploidy or <0.1% mosaicism in the fetus,) that all of the laboratories should biopsy all embryos, freeze them, and discard a lot of embryos before embryo transfer. Should government allow this kind of malpractice? Chinese Health Department has banned this malpractice already according to the common sense. Also in Australia, “the trial was suspended and then terminated early when we were unable to show an advantage for PGS” (Jansen, et al., 2008).

4. Basic principle of hereditary diseases is neglected in PGS. Heredity is the basic principle of the hereditary diseases. In chromosomal abnormalities and genetic diseases, patient's history are valuable source for tracing the possibility of the heredity of the offspring. PGT which is performed according to this tracing of patient's history will be very successful, because the imperfect check points and licensing system in these parents and families will be inherited to the embryos with very high possibility. However, PGS is to find out aneuploidy cells and omits the patient's history. More than 99% normal embryos which containing aneuploidy cells will not keep on with aneuploidy cells when they grow to be fetus (see above point 3), because those embryos inherit normal check points and licensing system. Thus the success rate and correct diagnosis rate of PGS is lower than throwing a coin (50%). Now there are 11 babies born from 26 mosaicism embryos. The results showed those mosaicism embryos contained zero (0 / 11) heredity to the babies. The PGS correct predictive efficiency for the babies is also zero (0 / 11). PGS designers cheat patients that the aneuploidy cells would be inherited to the offspring because DNA is hereditary material. They neglected that cytoplasm is also hereditary material. The check points and licensing system in the cytoplasm is controlling DNA replication and cell division. Without cell division, heredity of the aneuploidy cells will stop. These aneuploidy cells in the healthy embryos do not have characteristic of heredity. This is why the repeated same PGS results from day 3, day 5, day 12 to stem cells or to the baby born are very rare. And this is why so many “mosaicism” embryos can produce normal babies. 

In reverse, the embryos from the parents with history of hereditary diseases (such as Robertsonian translocation) will contain heredity characteristic. Some embryos inherit the parent’s imperfect check points and licensing system, which can not recognize the DNA mistake of Robertsonian translocation, and allow the abnormal cells keeping on the grow course to produce babies with the said hereditary diseases. Following the principle of heredity (patient's history) to perform PGD, the successful diagnosis rate is very high. In ESHRE PGD Consortium data from 2011 to 2012, 17721 PGD cycles were performed. It resulted in 3755 cycles with positive heartbeat. Only 34 misdiagnosis cases (0.9%) were reported (De Rycke, et al., 2017). It confirmed that PGD is 99% correct predictive efficiency as it is following patients’ family history, while PGS get <1% correct predictive efficiency as it is not following patients’ family history. They are significant different in statistics (chi-squared test p<0.0001). For the 45 year-old patients, PGS correct predictive efficiency may be higher but will not over 5%. According to statistics, if a clinical test can not success to 95% correct diagnosis rate, there will not be any meaning in clinical diagnosis. Thus, PGS is totally lack of any meaning in clinical diagnosis, and it is not qualified as a kind of diagnosis technique according to any international laboratory standard. PGS should be out of the clinical practice.

The failure of PGS also showed that genetics is out of its limit to explain: why sometimes some aneuploidy cells will lead to chromosomal abnormalities in babies (<1%) and why most of the aneuploidy cells (99%) found at the blastocyst stage by PGS do not contain heredity. It confirmed that Genetics is not the only theory to explain heredity and the control system of heredity. Evolution history showed: origin of heredity, i.e. origin of life, was not DNA, but a combination of RNA and lipid membrane. Whether the aneuploidy cells containing heredity characteristic or not, is depend on whether the check points and licensing system is normal or not. Whether the check points and licensing system is normal or not, can be simply according to the patient's history. Without patient's history, to screen aneuploidy cells in human embryos by PGS is aimless and is not scientific in diagnosis of hereditary problems. Hereditary diseases are combination results of both defects of DNA and heredity control system. To understand the heredity of cytoplasm and the heredity control systems in cytoplasm in human beings, especially in the embryo physiology, is imperative (please see attached paper “Origin of Life”.

When PGS does not contain predictive value in heredity, great scale of PGS contains the following malfunctions:

1. PGS is the biggest cheating event in reproductive field, which lasts about 25 years long, crossing two centuries, crossing many countries, with great scale of clinical malpractice on many patients.

2. PGS ruins and discards plenty of normal human embryos without any scientific reason.

3. PGS produces many unhealthy babies daily.

4. PGS will greatly increase government financial burden to support those unhealthy population such as Section 8 population in U.S.

5. PGS will change the normal human beings into subhuman beings in heredity.

Thus, PGS is a great scale of corruption case in U.S. and in the world.

In general IVF, some embryos are really aneuploidy, i.e. all of the cells in the embryos are all aneuploidy. However, most of them will arrest at any embryonic stages. They will not hatch and implant. Even after implantation, miscarriage will happen as natural selection. After the immune system in the fetuses set up, the normal fetal immune system will also stop the aneuploidy cells to inherit further. Only lesser than 1% baby will be born with chromosomal abnormalities. It shows that natural selection is much superior to artificial selection - PGS. PGS misled people to believe that most of the aneuploidy cells (DNA) found in the embryos are containing heredity characteristic.

5. Most of aneuploidy cells in early embryos have some normal and positive function in human embryos as a results of long evolution history.

A. They maintain the embryo internal environment, from maintaining stable temperature to transfer nutrition, to buffer outside mechanical stress, to keep the embryo spatial configuration and polarity before they go to apoptosis. They are not hyperplasia or malignant. It is foreseeable that when any embryo grows to 64 or 128 cell stage before implantation, some cells of the embryo will turn to be sick or aging (i.e. aneuploidy).  

B. Aneuploidy cells in trophectoderm contain more invasive strength than normal cells when puncture into maternal endometrium for implantation.  

C. Some aneuploidy cells in inner cell mass will continuously go to the course of apoptosis for organ and tissue differentiation. In the pig, it has been shown that the changed looser checkpoint system of spindle MAD-2 in oocytes leading to increased aneuploidy and apoptosis (Ma, et al., 2005). “The development of a healthy child requires not only very high rates of proliferation and differentiation, but also apoptosis, which is a crucial mechanism for morphogenesis and the development of the inner organs.” Changes in the regulation and rate of apoptosis in more or lesser level have deleterious effects on the trophoblast and consequently the developing embryo and fetus (Huppertz and Herrler, 2005). Without aneuploidy cells, without apoptosis, then without differentiation and without further healthy human babies. “In mammals, cellular proliferation, differentiation, and death accompany early placental development. Programmed cell death is a critical driving force behind organ sculpturing…” (Jurisicova, et al.,2005). In 83 human embryos biopsied on day 3 and analyzed on day 5, only one embryo (1%) consistently showed 100% normal 2N cells. However, that embryo was arrested on 12 cells on day 4 (Barrt, et al., 2004). It confirmed that: without aneuploidy cells, without apoptosis, the embryo could not grow further. The inference is: those rare blastocysts consistently containing 100% normal 2N cells are not heathy and can not survive long, due to lack of differentiation function at the time of differentiation. To call those normal embryos containing some aneuploidy cells at the time of fast differentiation to be abnormal and “mosaicism” is lack of scientific basis, and lack of common sense.

6. In 2008 ASRM meeting, Cooper Genomics (the biggest PGS company in the world) showed severe personnel quality problems. They hold a party meeting in the evening of Oct. 7th to talk about the techniques of next generation sequencing (NGS). In 30 minute talk, the speaker said five times that under a lot of condition, their diagnoses were subjective rather than objective. Usually three people gave three different diagnoses results while looking at the same DNA chart. It showed that even using the most advanced techniques could not solve their “aneuploidy” artifact. They joked that they were inventing PGT-AI (Artificial Intelligence) to solve the diagnoses problem. Their key mistakes are: the biopsied cell samples are full of DNA contamination by trophectoderm biopsy. No anyone of their team is responsible, and is with basic and full knowledge to design the PGT test from beginning to the end: such as from biopsy, to single cell amplification, primer selection, quality control set up, and clinical heredity possibility, etc. And their boss agreed about their severe weak points. The speaker showed their PGT-A (PGS) data (which is the largest collective data in the world for the time being, and most of embryo biopsy were performed by blastocyst biopsy): They have performed near to 5000 PGT-A cases with 60% pregnant rate and 50% birth rate. That is: PGT-A produced 10% (60%-50%) miscarriage rate. From year 2011 to 2015 CDC data, lesser than 35 years old group, the miscarriage rate were all about 7% (pregnant rate minus birth rate). This confirmed that PGT-A (PGS) increased rather than decreased the miscarriage rate in great scale clinical practice. It meant that PGT-A increased aneuploidy rate after trophectoderm biopsy, laser radiation, and freezing-thawed procedures. The reason is:

Part II: PGT is not safe

1. Great scale of PGS without patient's history is a great scale of holocaust of “Eugenics”. If 60% of human embryos were diagnosed as aneuploidy or mosaicism, in which identical repeated results to be (13/53) 24.5% (Munné, et al. 2005), the misdiagnosed and discarded (or killed) embryos will be 60% X (1 - 24.5%) = 45.3%. The Society for Assisted Reproductive Technology (SART) data of all age groups in 2016 showed IVF cycles to be 115,610. If average embryos for PGT-A per cycle to be five, and only half of the patients to be performed with PGT-A, total embryos to be discarded was 5 X ½ X 115,610 X 45.3% = 130,928. If 30% of these embryos could be produced to be babies, and year 2017 and 2018 IVF data to be the same as 2016, total number of embryo to baby life to be killed (from 2016 to 2018) would be 130,928 X 30% X 3 = 117,835. It is not safe, not humanity and not ethical. Those several notorious repeated promoters for PGS should take responsibility of this large scale of cruel and shameful holocaust in 21 century. Is it safe? Can any country allow this kind of embryonic killing (“Eugenics”) to be continuing?

2. The following factors will lead microtubules and other cellular organelles to be damaged, thus they will produce aneuploidy cells in our embryos at any time:

A. Environment factors: room and culture temperature change, the embryos moving out incubator repeatedly, not suitable culture medium, not enough nutrition, bad air, etc.

B. Foreign and mechanical strength: suction strength, denude strength, biopsy pull and push strength, etc.

C. Physical factor: laser (electromagnetic radiation), freezing ice expansion, freezing and thawing osmolarity change, etc.

D. Chemical factor: cryoprotectant DMSO, HCl acid, etc.

To use aneuploidy-producing techniques to prevent aneuploidy is a joke. It is not scientific.

3. Embryo biopsy is traumatic, and is not safe for most manipulators. Embryo biopsy is safe were first time systematically analysed and announced with mice model under specific and strict conditions rather than random condition (Cui, et al., 1993a, 1993b, 1994). The “embryo biopsy is safe” a relative speaking. The papers confirmed that embryo biopsy will not reduce the cell number in the offspring body, organs and tissues. They also confirmed that embryo biopsy will not produce deformed organs or tissues from heart, kidney to bone skeleton, etc. The papers also confirmed that the function of liver, kidney and bone marrow (blood-making function), etc. will not be harmed by embryo biopsy. However, the author also set up the condition of safety for embryo biopsy: First, stage for embryo biopsy: “Embryo biopsy at the 8-cell stage may therefore be the optimal choice for preimplantation diagnoses”, rather than trophectoderm biopsy (Cui, 1993 Page 14-16). Second condition of safety for embryo biopsy was set up according to the results of 13 series biopsy experiments which showed how to improve the skill and technique of biopsy (Cui, et al. 1993c). The condition is: the hatching rate (rather than blastocyst rate) after embryo biopsy should be more than 90% as a simple and reliable guide to evaluate the technical skill of single cell embryo biopsy. Such high hatching rate will result in equivalent rates of implantation and fetal development for both biopsied and non-biopsied control groups. (Cui, et al. 1993c, Cui, 1993, Page 42-55). This condition of safety for embryo biopsy (i.e. 90% hatching rate after day 3 embryo biopsy) is too high demand for most of laboratory staff to succeed in this most difficult micro-manipulation. Most papers published about the results of embryo biopsy did not show to be the same results as the control group in embryo development, implantation rate, fetus rate and normal birth rate. Thus, embryo biopsy is not safe (in most IVF laboratory staff’s hands) is an absolute speaking. Excellent embryo biopsy performance needs strong basis of Embryology theory, very good brain-hand coordination, very patient temper and very responsible characteristic for the health of our offspring with rich practical training. In human embryo biopsy (biopsy using acid, using laser or using mechanical cutting to open the human zona pellucida), the only best and safest technique is day 3 biopsy with mechanical zona cutting (one point to three-dimensional cutting), which has not yet been popularized. The opened zona location by this cutting is like a blooming flower. Smooth bevelled needle will be easily pointed to the selected aim cell for biopsy. If using other unsafe biopsy techniques, some remained embryonic cells will be pushed or pulled to be deformation during embryo biopsy, which will influence microtubules in the cells to be broken or to be dislocated. The results are: the later differentiation of neuron, immune system, and heart, other organs and tissues will be abnormal functioning (See attached paper “Origin of Life”, part of differentiation). The offspring will be not healthy, with inferior function of their organs and tissues, especially with obvious mental problems. They are not normal human beings but subhuman beings. How healthy or how clever the future babies will be depends on the laboratory staff’s techniques, skills and their time and responsibility during manipulations. Thus embryo biopsy should not be performed in great scale as the recent PGS or PGT-A. Embryo biopsy should never be recognized as a safe technique for most staff as popular usage, and the PGT patients should be only limited to families with hereditary diseases.

When embryo biopsy is performed by inappropriate workers under loose condition: wrong stage, wrong techniques and bad skills, such as lesser than 90% hatching rate or not care hatching rate, embryo biopsy will really produce unsafe results. How to differentiate safe or unsafe results in human embryo biopsy? They are described as the following:

A. After embryo biopsy, the biopsied embryos will delay several hours to allow the check point and licensing system (see attached paper –“Origin of Life”) to restore their function to be normal. Then cell cleavage will goes on. The delay time for one point to three-dimensional mechanical cutting, acid cutting and laser cutting are each about 2 hours, 3 hours and 4 hours. The laser cutting will delay the longest time. If embryo biopsy harm microtubules, other cytoskeleton and DNA replication, which influence centrosome function, those cells will arrest. No cleavage will happen in those cells. If some of microtubules to be harmed by biopsy, some cells will cleave with abnormal chromosomal condition, aneuploidy cells will be produced.

B. After embryo biopsy, the biopsied embryos will grow to expanded blastocyst. The embryos performed by one point to three-dimensional mechanical cutting will arrive this condition as the non-biopsied ones at the same time, while laser cutting will delay 2-4 hours to arrive this condition. Only under very poor embryo biopsy condition, the blastocyst rate after biopsy will be lower than the control group. The blastocyst rate is not a sensitive indicator to evaluate the biopsy skill and technique (Cui, et al., 1993c).

C. The good biopsied embryos will easily arrive hatching and hatched stage to get 90% hatching rate.

D. The good biopsied embryos will achieve the same implantation rate as the non-biopsied ones.

E. The good biopsied embryos will achieve the same fetus rate as the non-biopsied ones.

F. The good biopsied embryos will achieve the low miscarrage rate as the non-biopsied ones. If the biopsied embryos get too many aneuploidy cells to influence their normal functions, they will be miscarried.

G. If the microtubules and actins of the cells are harmed by embryo biopsy which influence the normal differentiation of the fetal heart muscles, the fetal nervous system and endocrine system, stillbirth will happen (See attached paper – “Origin of Life”).

H. After birth, the unhealthy offspring may show different symptoms and signs, which may be simply called embryo biopsy syndrome. The unhealthy offspring data in mice after blastomere biopsy were concluded by Zacchini et al. as: increased or decreased body weight, impaired memory, impaired locomotor coordination, increased anxiety, altered response to stress, structural abnormalities in the corpora striatum and hippocampus, epigenetic and proteomic defects in brain, impaired function of adrenal glands, and reduced fertility in 40-week-old females (Zacchini, et al., 2017). According to these scared and severely unhealthy data, almost all of PGT centers in U.S.A. stopped blastomere biopsy, and remarkably switched to blastocyst biopsy since 2015 with laser cutting, and freezing all biopsied embryos before next time for embryo transfer. However, all of the papers Zacchini et al. reviewed and quoted (about the offspring results) were all mistakenly using very harmful chemical - cytochalasin B in the biopsy medium in very high concentration, (Yu, et al., 2009; Zhao, et al., 2013; Zeng, et al., 2013; Yu, et al., 2013; Sampino, et al., 2014; Wu, et al., 2014). None of those papers showed the manipulators got high hatching rate after blastomere biopsy, to confirm their techniques to be qualified for embryo biopsy. They also selected the harmful 4-cell stage to manipulate the embryos. Cytochalasin B was known as important microtubule and microfilament-disrupting drugs (Gregoraszczuk and Stoklosowa, 1997; Martin, et al., 1981). When microtubule and microfilament are disrupted, all kinds of abnormalities of organs and tissues will be produced. To use this kind of animal experiments to give a warning, to the field of human reproduction medicine not to perform embryo biopsy randomly as PGS, may be helpful. Nevertheless, the effect of pushing embryo biopsy from 8-cell stage to blastocyst biopsy is misleading. The popularized blastocyst biopsy for PGS for every IVF embryos is much more misleading. It is leading to the direction of subhuman reproduction with plenty of offspring, with mental and other problems, to be produced daily, monthly and yearly.