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PASSADO
470 a 402 AC: DEMCRITO SUGERIU QUE O TESTCULO DIREITO PRODUZIA MACHOS E O ESQUERDO FMEAS. DVIDA PERMANECEU AT O PASSADO RECENTE
09/12/2010
XY Then Achieved:
XY Inc.
dlg
SX MoFlo
Separacin de espermatozoides
3. Los zoides X fluorescen con 4% ms de intensidad que los Y 4. La computadora procesa la fluorescencia detecta-da y categoriza los zoides como X, Y o dudosos 6. Las gotculas cargadas son desviadas cuando pasan frente a placas continuamente cargadas
1. Un
cristal piezoelctrico ondula y quiebra la corriente en gotculas ~ 90,000/second. 2. Un rayo laser proyecta luz azul sobre los zoides
5. Se aplica una carga negativa, positiva o ninguna a las gotculas 7. Las muestras son recojidas en 3 recipientes
XY, Inc.
Multiplicacin de
de semen sexado
Pietro S. Baruselli
XY Inc.
dlg
09/12/2010
Roteiro da apresentao:
Inseminao artificial com smen sexado
Hiptese experimental:
A reduo no intervalo entre a inseminao e a ovulao proporciona aumento na taxa de concepo em vacas inseminadas com smen sexado
Ejaculado
Congelamento convencional
Sexagem
Congelamento
eCG (400UI)
Norgestomet (Crestar)
D0 D9
48h
eCG (400UI)
Norgestomet (Crestar)
D0 D9
48h
Figura. Efeito do momento da IA (54h ou 60h aps a retirada do implante de progestgeno) e do tipo de smen (convencional ou sexado) na taxa de concepo aps IATF em vacas Nelore (Bos indicus) no ps-parto.
Souza et al., 2007
09/12/2010
60
a
50
60 50
P > 0.10
Taxa de concepo
Taxa de concepo
n = 193
40
b
n = 196
n = 195
40
n = 194
30 20 10
30
20
10
Convencional
Sexado
IATF 60h
IATF 54h
Grupos experimentais
Grupos experimentais
Research colaboration USP and Sexing Dr. Nil Campos Filho PRELIMINARY RESULTS USA EXPERIMENTS
EXPERIMENT 1
09/12/2010
OBJECTIVE
The aim of this study was to evaluate the follicular dynamics in Jersey heifers (Bos taurus) submitted the FTAI.
EXPERIMENTAL DESIGN
EB + PGF
PGF
EB
FTIA
US 0 hours
US 24 hours
US 48 hours
US 60 hours
US 65 hours
Table - Follicular dynamics in Jersey heifers (Bos taurus) submitted the FTAI.
Animals number 26
92.3 (24/26) 66.12.2 16.02.3 15.62.3
RESULTS
Ovulation rate (%) Interval device removal/ovulation (h) > Dominant Follicle (mm) > Ovulatory Follicle (mm)
100 90 80
Percentage of ovulation
88
EXPERIMENT 2
70 60 50 40 30 20 10 0 48 60 66
Hours after device removal
6 0
6 0 72 84
Figure - Time of ovulation in Jersey heifers (Bos taurus) submitted the FTAI.
09/12/2010
progesterone device removal and artificial insemination on pregnancy rates in Bos taurus (Jersey) heifers fixed time inseminated with sexed or non-sexed semen.
We hypothesized that delay timed AI (TAI) with sexed semen increase the pregnancy rates in Bos Taurus (Jersey) heifers.
EXPERIMENTALS DESIGN
2 x 2 Fatorial: (type of semen vs. timed AI) Groups: 1) Non-sexed semen 54h - AI 12h before ovulation. 2) Non-sexed semen 60h - AI 6h before ovulation. 3) Sexed semen 54h - AI 12h before ovulation. 4) Sexed semen 60h - AI 6h before ovulation.
D0
TREATMENT
1) Group FTAI 54h
FTAI Sexed or Non-sexed EB SEMEN (1.0mg) EB + PGF (2mg) (12.5mg) PGF (12.5mg)
CIDR
D0 D8
54h
D9 D10 D12
CIDR
D8
60h
D9 D10 D12
P=0.001
51.2
RESULTS
40
23.7
30
n=213
20
n=207
10 0 Non-sexed Sexed
Figure 1 - Conception rate by semen type (Non-sexed and sexed) in Jersey heifers submitted to FTAI.
09/12/2010
P=0.047
50.5a
51.8a
40 30
20 10 0
n=101
31.4b
n=112
16.2c
n=105
n=102
20 10 0
n=206
Figure 2 - Conception rate by timed AI (54h vs 60h after withdrawal P4 device) in Jersey heifers submitted to FTAI.
Figure 3 - Conception rate by semen type (Non-sexed and sexed) and timed AI (54h vs 60h) in Jersey heifers submitted to FTAI.
P=0.001
60
44.2a
41.3a
Pregnancy rate (%)
50 40
47,2
ab
30,9
30 20 n=71 10 0 n=68
ab
35,2
23,9
n=64
n=74
n=71
n=72
54H
60H
54H
60H
54H
60H
755
760
762
Bull 755
Bull 760
Bulll 762
Figure 4 - Conception rate by bull (755, 760 and 762) in Jersey heifers submitted to FTAI.
Figure 5 - Conception rate by bull (755, 760 and 762) and timed AI (54 and 60h) in Jersey heifers submitted to FTAI.
100 90 80
Pregnancy rate (%) 100 90 a ab
70 60 50 40
bc
61,4
Pregnancy rate (%)
80 70 60 50 40 30
n=35
52,8 39,4
e
n=71 n=72 n=66
cd
34,9
n=70
ed
38,9 25,8
30
20 10 0
Non-sexed Sexed
21,9
n=73
n=33 n=35
28,6 15,8
n=38 n=35
14,7
n=68
21,9
n=36
n=33
n=39
20 10
n=31
8,3
n=36
n=32
Non-sexed
Sexed
Non-sexed
Sexed
0
54H 60H 54H 60H 54H 60H 54H 60H 54H 60H 54H 60H
755 Non-sexed 755 Sexed 760 Non-sexed 760 Sexed 762 Non-sexed 762 Sexed
Bull 755
Bull 760
Bulll 762
Figure 6 - Conception rate by bull (755, 760 and 762) and semen (Non-sexed and sexed) in Jersey heifers submitted to FTAI.
Figure 7 - Conception rate by bull (755, 760 and 762), semen (Non-sexed and sexed) and time of FTAI (54 and 60h) in Jersey heifers.
09/12/2010
CONCLUSION
Jersey heifers inseminated with sexed semen showed lower pregnancy rate than non-sexed semen. However, pregnancy rates with sexed semen improved with the further increase of 4h in the interval between the P4 device withdrawal and the FTAI, confirming our initial hypothesis. The 755 bull showed lower pregnancy rate than the 760 and 762 bulls.
EXPERIMENT 3
OBJECTIVE
Rate and time to ovulation in Jersey
The aim of this study was to evaluate follicular
dynamics in Jersey heifers (Bos taurus) synchronized with new and previously used progesterone device in FTAI protocols.
EXPERIMENTAL DESIGN
EB + PGF NEW or USED CIDR D0 D8 D9 US 24/24h D0 D8 D9 D10 D10 US 12/12h D12 D12 PGF EB FTIA
RESULTS
US 0 hours
US 24 hours
US 48 hours
US 64 hours
09/12/2010
Table - Follicular dynamics in Jersey heifers (Bos taurus) synchronized with new and used device progesterone in protocols to timed AI.
Percentage of ovulation (%)
New
100 90 80 70 72 64
Used
CIDR P New Animals number Ovulation rate (%) Interval device removal/ovulation (h) > Follicle Dominant (mm) > Follicle Ovulatory (mm) 27 92.6 (25/27) 66.90.5 15.50.5 15.10.5 Used 21 100.0 (21/21) 65.5 0.7 14.3 0.5 14.2 0.5 0.98 0.24 0.18 0.33
60
50 40 30 20 10 0 48 60 65 Hours after device removal 72 84 0 0 4 8 20 12 4 0
Figure - Interval between P4 device removal and ovulation in Jersey heifers synchronized with new and previously used progesterone device in FTAI protocols
CONCLUSION
We conclude that new and used device of progesterone presented similar rate and timing
EXPERIMENT 4
We hypothesized that delay timed AI (TAI) with sexed semen increase the pregnancy rates in Bos taurus (Jersey) heifers.
09/12/2010
EXPERIMENTALS DESIGN
2 x 2 Fatorial: (Timed FTAI vs type of CIDR) Groups:
TREATMENT
Groups FTAI 50h /New or used CIDR
EB + PGF (2mg) (12.5mg) PGF (12.5mg) EB (1.0mg) FTAI
50h
D8 D9 D10 D12
1) New CIDR and FTAI 50h. 2) Used CIDR and FTAI 50h. 3) New CIDR and FTAI 56h. 4) Used CIDR and FTAI 56h.
D0
56h
D9 D10 D12
P=0.18
50
45 40 Pregnancy rate (%) 35 30 25 20 15 10 5 0 50h 56h Hours after removal device
n=208 n=200
RESULTS
19.5
14.9
Figure 1 - Conception rate by timed AI (50h vs 56h after removal P4 device) in Jersey heifers submitted to FTAI.
P=0.28
50 50 45 45 40
P=0.28
35 30 25 20 15 10 5 0 New CIDR - FTAI New CIDR - FTAI Used CIDR - FTAI Used CIDR - FTAI 50h 56h 50h 56h n=107 n=99 n=101 n=101 13.1 18.2 20.8 16.8
18.8 15.5
New CIDR
Used CIDR
Figure 2 - Conception rate by progesterone device (new vs 1x used) in Jersey heifers submitted to FTAI.
Figure 3 - Conception rate by bull (755, 760 and 762) in Jersey heifers submitted to FTAI.
10
09/12/2010
50
50 45 40 Pregnancy rate (%) 35 30 25 20 15 10 5 0
8.5
n=47 n=25 n=19 n=82 n=72
45 40
34.2 28.6 24 19.1
n=79
34.2
10.5
12.2 8.3
n=7
10.3
n=21 n=29
10 7.1
n=10 n=17
492
503
504
753
760
761
762
763
766
767
773
492
753
760
761
762
50 45
40
Pregnancy rate (%) 35 30 25 20 15 10 5 0 FTAI 50h FTAI 54h
n=208 n=105
Experiment 2 Experiment 4
31,4
FTAI 56h
FTAI 60h
Figure 6 - Conception rate by timed AI (50h, 54h, 56h and 60h after withdrawal P4 device) in Jersey heifers submitted to FTAI.
Sexed Unsexed
Holstein
Red Breed
Jersey
Taxa de prenhez de vacas Nelore inseminadas com smen sexado conforme o touro (n=3.200 IA), Fazenda Campo Verde e Paraso, Viana-ES, 2006/2007
11
09/12/2010
EXPERIMENT 5
Holstein (-12,6%) Red Breed (-7,3%) Jersey (-5,2%)
Efeito de raa ?
OBJECTIVE
Rate and time to ovulation in Jersey
The aim of this study was to evaluate the follicular dynamics in Jersey heifers (Bos taurus) in normal estrous cycle.
ULTRA-SONOGRAPHY EXAMS
ESTROUS US US US US
D0 AM
D0 PM
D1 AM
D1 PM
D2 AM
RESULTS
US D0 AM
US D0 PM
US D1 AM
12
09/12/2010
Table - Follicular dynamics in Jersey heifers (Bos taurus) in normal estrous cycle.
Animals number Ovulation Rate until 48h (%) Interval ovulation: Onset heat (h) End heat (h) > Follicle Ovulatory (mm) 31.2 0.9 16.3 1.2 14.1 0.3 43 86.1 (37/43)
Percentage of ovulation
32,4
29,7
27,0
10,8
n=04
n=12
n=11
n=10
Figure - Heat onset to ovulation interval in Jersey heifers (Bos taurus ) identified by Heat Watch system.
EXPERIMENT 6
Timing of insemination with sexed or nonsexed semen on pregnancy rates of Jersey heifers identified in estrus by radiotelemetric estrus detection system
TREATMENT
2 x 4 Fatorial: (type of semen vs timed AI)
Groups:
a) Sexed semen 1) AI 0 to 6 hours after onset heat 2) AI 6 to 12 hours after onset heat 3) AI 12 to 18 hours after onset heat 4) AI 18 to 24 hours after onset heat b) Non-sexed semen
EXPERIMENTALS DESIGN
AI Sexed or Non-sexed SEMEN Heat Watch + PGF
Heat detection for Heat Watch
US
1)
2) 3) 4)
13
09/12/2010
P=0.001
100 90 80
RESULTS
a 64,2 b 49,5
n= 371 n= 382
Sexed
Non-sexed
Figure 1 - Conception rate by semen type (Non-sexed and sexed) in Jersey heifers identified in estrus by Heat Watch system.
Sexed
80
70
Non-sexed
70
60
65,5
60 50 40
n=60 46,9 47,2
58 53,6
n=77 n=72
n=84 n=57
n=58
30
n=64
20 10 0
0 to 5 5 to 10 10 to 15 15 to 20 20 to 25
Figure 4 - Timing of insemination with sexed or non-sexed semen on pregnancy rates of Jersey heifers identified in estrus by Heat Watch system.
755 BULL
Sexed
80 70 60 Pregancy rate (%) 48,2 50 40 30
n=85 n=78 n=111 n=104 n=99 n=100 n=84 n=88
Non-sexed
Sexed
68,1
Non-sexed
62,8 48,7
64,7
61,1
0 to 6
6 to 12
12 to 18
18 to 24
Figure 5 - Timing of insemination with sexed or non-sexed semen on pregnancy rates of Jersey heifers identified in estrus by Heat Watch system.
Figure 6 - Timing of insemination with sexed or non-sexed semen on pregnancy rates of Jersey heifers identified in estrus by Heat Watch system.
14
09/12/2010
760 BULL
Sexed 80 70 60 Pregancy rate (%) 50 40 30 20 10 0 0 to 6 6 to 12 12 to 18 18 to 24 Interval from first standing event to AI (h)
n=40 n=38 n=51 n=42 n=36 n=43 n=41 n=42
762 BULL
Sexed
65,9 69,1 80 70 60 Pregancy rate (%) 59,1 51,6 44,1 27,6
n=22 n=25 n=29 n=31 n=35 n=30 n=19 n=24
Non-sexed 74,4
Non-sexed
70,1 58,3 48,5 36,8
50 40 30 20 10 0
0 to 6
6 to 12
12 to 18
18 to 24
Figure 7 - Timing of insemination with sexed or non-sexed semen on pregnancy rates of Jersey heifers identified in estrus by Heat Watch system.
Figure 8 - Timing of insemination with sexed or non-sexed semen on pregnancy rates of Jersey heifers identified in estrus by Heat Watch system.
Sexed
70 60,1 60 48,8 Pregancy rate (%) 50 40 30 20 10 0 > -8 -8 to 0
n=86 n=70 n=110 n=120
Non-sexed
66,7 63,7 51,5 63,3
48,2
48,5
n=60
0 to 8
>8
Figure 9 - Timing of insemination with sexed or non-sexed semen on pregnancy rates of Jersey heifers identified in estrus by Heat Watch system.
Sexed
70 59,8 60 Pregancy rate (%) 50 40 30 20 10 0 0 to 10 10 to 20 20 to 30
n=91
Non-sexed
64 ab 50,7 a 56,1 ab 45,8 66,7 67,2
a 57,8
64,5
NUMBER OF MOUNT
b 38,5
n=92 n=90
n=76 n=77
n=75 n=50
n=63 n=59
n=58
30 to 40
> 40
Figure 10 - Conception rate by number of mount in Jersey heifers identified in estrus by Heat Watch system.
15
09/12/2010
Sexed
70 60 Pregancy rate (%) 50 40 30 20 10 0
n=55
Non-sexed
66,7 ab a 50,9 67,6 a 62 ab ab 46,2
b 54,1
a 57,1
DURATION OF HEAT
b 34,6
n=61 n=84 N=81 n=177 n=151 n=52 n=71
0 to 6
6 to 12
12 to 18
18 to 24
Figure 11 - Conception rate by duration of estrus in Jersey heifers identified in estrus by radiotelemetric estrus detection system .
CONCLUSION
Our initial hypothesis that AI using sexed semen performed around the end of the estrus
Acknowledgements
Sexing Technologies and Dalhart Jersey Ranch
increase
confirmed.
the
pregnancy
rates
was
not
Next studies using more animals should be conduced to confirm this preliminary results.
INSEMINACION ARTIFICIAL A TIEMPO FIJO PROFUNDA CON SEMEN SEXADO EN VAQUILLONAS HOLANDO
G. M. Brogliatti, G. Domnguez, M. G. Lusenhofft, J. Perkins, G. A. Bo, Cutaia L.
1Centro de Inseminacion Artificial La Argentina Chica, Vdo Tto; 2 Actividad Privada Venado Tuerto; 3 Adeco SA; 4 IRAC Instituto de Reproduccin Animal de Cordoba,
Introduccion
Poder elegir el sexo en los mamferos ha sido uno de los objetivos ms buscados durante mucho tiempo por los cientficos y ganaderos del mundo.
16
09/12/2010
Test Termo-Resistencia
Porcentaje de espermatozoides mtiles progresivos
Trabajos previos
Lugar de Inseminacion Seidel y Col. CSU (1999)
Horn 27/49 68% Body 25/50 60% 85/158 53% 70/163 40%
0 (%) 23,14,9
Trabajos previos
Horario de la IATF Veneranda y Col
52 Horas 18/50 60 Horas 20/50 54 Horas 72/185 36 % 40 % 37 %
Objetivo
Comparar los porcentajes de preez con semen sexado (2 M de esp.)
Depositados en Cuerpo (tradicional) vs Cuerno (profundo) 52 h vs 58 h posterior al retiro del dispositivo intravaginal.
Capitaine
1241 servicios en 17 establecimientos en vaquillonas, 37 % (34-40 %)
17
09/12/2010
Materiales y Metodos
Vaquillonas Holando Argentino entre 15 y 18 m
condicin corporal 3,05 + 0,22. evaluadas por ecografa para determinar estructuras ovricas y tamao del tero inicio
Materiales y Metodos
Un solo inseminador con una jeringa tradicional para IA en el cuerpo y la jeringa de TE para las inseminaciones en el cuerno uterino (vainas de ET, SBS Cryotec sa, Arg.). Diagnstico de preez ecografia transrectal 30 d Analizados por regresion logistica y al combinar los datos por Chi Cuadrado (Infostat).
Al momento de la IA, ecografa para determinar el tamao y el lugar del folculo preovulatorio.
Resultados I
52 hs
Cuerno Cuerpo 21/43 49 % 17/45 38 % 38/88 43 %
58 hs
15/24 63 % 21/51 41 % 36/75 48 %
Total
36/67 53 %a 38/96 39% b
PG
IATF 52
9
B.E
CIDR (8 d)
8 10
PG
PG
9
B.E
10
Total
58 hs
32/65 49,2 % 35/104 33,6 % 67/169 39 %
Total
64/155 41 % 70/202 34 % Vaq. 357
Total
18
09/12/2010
Ejaculado
Retirada do CRESTAR
PGF2
BE (2mg)
Colheita de Embrio
D0
D4
D5
D6
D7 T
D8 M
D8 T
D9 M
D15 M
Congelamento convencional
Sexagem
BE (2mg)
Retirada do CRESTAR
Colheita de Embrio
Congelamento
D0
D4
D5
D6
D7 T
D8 M
D8 T
D9 M
D15 M
Experimental design
1st Replicate
Smen convencional (n=5)
(35d)
2nd Replicate
Smen sexado (n=5)
Tabela 1 - Produo de embries em vacas Nelore (Bos indicus) superestimuladas e inseminadas em tempo fixo com smen sexado e convencional, Poos Caldas, MG, 2007.
Smen Convencional (n=10)
Total estruturas (n) Emb. transferveis (n) Emb. congelveis (n) 9.90 0.78 6.80 0.66 5.90 0.71 1.50 0.48 1.60 0.37 68.7
Emb. no fertilizados (n) Emb. degenerados (n) Taxa de emb. viveis (%)*
59.6
41.7
0.02
0.32
0.73
Experimental design
1st Replicate Group Conv 12/24h (n=4)
D15 AM (60d)
2nd Replicate Group Sexed 12/24h (n=4) Group Sexed 18/30h (n=4) Group Conv 12/24h (n=4)
(60d)
3rd Replicate Group Sexed 18/30h (n=4) Group Conv 12/24h (n=4) Group Sexed 12/24h (n=4)
AI
AI
12h
D8 PM
24h
D9 AM
DIB removal
Flushing
AI
AI
18h
30h
D8 PM D9 AM
D15 AM
Group Convencional semen (AI 12/24h after LH) Group sexed semen (AI 12/24h after LH) Group sexed semen (AI 18/30h after LH)
19
09/12/2010
8 7 6 5 4 3 2 1 0
Mdico Veterinrio: Claudiney de Melo Martins Fazenda: Santa Brbara, Botenhos - MG
Transferable embryos
Conv 12/24h
Sexed 12/24h
Sexed 18/30h
Figure - Effect of type of semen (conventional vs. sexed) and timed artificial insemination after LH (12 and 24 vs. 18 and 30) in Nelore cows (Bos indicus) superstimulated, Poos de Caldas, MG, 2008. Martins et al., 2008
AI
AI
12h
D8 PM
24h
D9 AM
D15 AM
DIB removal
PGF2
LH 24h FSH (200mg
Folltr.)
Flushing
9 8 7 6 5 4 3 2 1 0
8.92
a
5.92 4.41
b
Transferable embryos
Conv 12/24h
Sexed 12/24h
Sexed 18/30h
AI
AI
18h
30h
D8 PM D9 AM
D15 AM
Figure - Effect of type of semen (conventional vs. sexed) and timed artificial insemination after LH (12 and 24 vs. 18 and 30) in Holstein heifers (Bos taurus) superstimulated, Poos de Caldas, MG, 2008. Martins et al., 2008
Retirada do CRESTAR
PGF2
BE (2mg)
Colheita de Embrio
D0
D4
D5
D6
D7 T
D8 M
D8 T
D9 M
D15 M
Retirada do CRESTAR
PGF2
BE (2mg)
Colheita de Embrio
D0
D4
D5
D6
D7 T
D8 M
D8 T
D9 M
D15 M
20
09/12/2010
INTRODUCCION OBJETIVOS
MATERIALES Y METODOS
PROTOCOLO DONANTES CON DISPOSITIVO Dia 0 8.00 h. Colocacion Disp.+p4 + 17bEstradiol Dia 5 8.00 h. 3cc Folltropin V 18.00 h. 3cc Folltropin V Dia 6 8.00 h. 2cc Folltropin V 18.00 h. 2cc Folltropin V Dia 7 8.00 h. 1.5cc Folltropin V+ 25mg dinoprost 18.00 h. 1.5cc Folltropin V+ Ret. Disposit. Dia 8 8.00 h. 1-0.5cc Folltropin V 18.00 h. 1-0.5cc Folltropin V Dia 9 Control de celo + IA Dia 16 Lavaje y Transf
Dia15
Dia 16 Dia 17
8.00 h.
18.00 h. 8.00 h. 18.00 h.
Dia 24
Lavaje y Transf
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09/12/2010
PRODUCCION DE EMBRIONES MEDIAS +- EE EN VACAS ANGUS SUPEROVULADAS E INSEMINADAS CON SEMEN SEXADO O CONVENCIONAL
PRODUCCION DE EMBRIONES MEDIAS +- EE EN VACAS ANGUS SUPEROVULADAS E INSEMINADAS CON SEMEN SEXADO O CONVENCIONAL
SEMEN SEXADO Toro Rp N de donantes inseminadas Ovocitos/ embriones totales Embriones fertilizados Embriones transferibles % Embrionestransferibles /ovocitos embriones totales Transferencias en fresco Preadas % Preez en fresco RP 251 29 13+2,5a 9+1,6 7,3+1ab 60a 32 19 59.4a RP 253 5 17.3+4,9a 11,7+2,9 10,2+2,4b 51.6a 20 13 65a RP 913 10 20.3+7,5 a 3,4+1,2 2,8+1a 15.6b 10 3 30a TOTAL 44 15.51+2,3 a 8,4+1,3a 6,94+1a 41.69a 62 35 56.5a SEMEN NO SEXADO 27 10.5+ 1,5a 7,6+1,2a 6,83+ 1,16a 64.43b 65 25 38.5b
NO SEXADO Tratamientos Superovulatorios Ovocitos/embriones totales Embriones Fertilizados Emb Transferibles/Vaca 29 10.5+ 1,55 a 7,62+ 1,25 a 6,83+ 1,16 a
% EMB. / ovocitos embriones totales Transferencias en fresco Preadas(Transf. En fresco) % Preez en fresco
64.4a 65 25 38.5a
52.8b 52 32 61.5b
Cometarios
Comentarios Generales
Muchas Gracias
22