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- f: Ith Orqanization fi].:TAcH vA PHAN LAp ::t::;~l~ *~-~-, .,, :' e perinatal anc ~;: TAP CHi Y - D110C HOC QUAN Sl1 s6 6-2014 ~D"'"": - TE BAo PHOI THAI TRONG MAu NGOAI VI -'" "" "" It CUA ME UNG DUNG CHAN £lOAN KHONG CAN THIEP • • • f?:'::>"c' determinants ir ;:.,10" Tri~u tu« Sang*; Tr~n Van Khoa*; Dinh Doim Long** 'aediatr Perinat;;;:'?:'H)M ;-545 'Nam 1893, Schmorl phat hien t9 bao (TB) ph6i thai & ph6i cua phu n(Y mang thai Toi nay, , nhi~u nghi~n cou, oa kh~ng ojnh Sl,l' c6 mat cua TB ph6i thai rnau ngokit s MultiVysion PGT Multicolor probes, kinh, lamen, NP40, SSC20X, antifade II, Ii ky, kh6 ide, nU'&c tinh khi~t, HCI, methanol, hi$u; ho )n tal tro b~ mat d~ Thitit b] may moe Cl,I th~ cac buoc nhu sau: + BU'ac 1: 11Iy10 1-11khanq h6a vao 6ng 1,5 ml th~ da biotin + Buoc 2: them 20 1-11 hat tlr da gan streptavidin, Ll (y 4°C 30 phut + BU'ac 3: them 100 fJl PBS 1X, lac nhe, rem th~ g; May I~c 6n nhi$t (Eppendorf, f)(rc), may diu nhar tam cao t6c (Hettich, f)(rc), pipet man i thai trorl~lf;i:tGlnson, My), buong thao tac PCR (My), rO'n,;:i.t:~,e;a ttY tinh (Promega, My), b~ nuoc 6n nhiet , h$ th6ng kinh hi~n vi huynh quang io Zeiss, f)(rc) PhlPO'ngphap nghien ciJ'u Quy trlnh thl,l'c hi$n d~t vao gia tlr tinh, hut be dich + BU'ac 4: them 100 fJl PBS 1X, lac nhe, hut h~t djch cho vao 6ng chua mau ngo?i vi cua me da lay tnroc Lac quay tron 15 phut (y 4°C + BU'&c 5: hut tlrng ml mau cho vao ong 1,5 ml f)~t len gia tLl' tinh hut be toan b(> djch, giCr I?i ph(J'c hQ'p h?t tLl' - hong c~u nhan da bi g~n I€m tren bng phia gia ttY tlnh + BU'&c 6: them 100 fJl PBS, lac d~u, T90 phCtc hc '0 nhan TB BU'ac 3: nia lam kfnh b~ng dung dich PBS 1X (pH 7,0: 0,14 M NaCI, mM KCf, 10 mM Na2HP04, mM KH2P04) phut (y nhi$t dO phong RLra s?ch nLl'ac cllt v6 trung I~n Lo?i nLl'ac b~ng dung dich 70%, 90%, 100% phut (y nhi$t d(>phong va lam kh6 kh6ng 51 56 6-2014 TAP CHi Y - DU'OC HOC QUAN SU' BU'ac 4: them j.J1hen hop probe va phu len tren b~ng lamen kich thuo'c x mm Bao phu xung quanh b~ng lap nhva g~n cao 5U d~ tranh thoat hO'i ruroc BU'ac 5: u block nhi$t 75°C phut, sau lai tLi' 16 - 22 gia tIm tu BU'ac 7: nra nghieng mat lam kinh bang dung dich O,4X SSC/O,3% NP40 & 73°C phut BU'ac 8: rLra lam SSC 2X BU'ac klnh dung die!", & nhlet dQ ph6ng phut 9: lay lam kinh d~t phon~ t6i, dung giay m~m lau kho Them ~ 37°C BU'ac 6: tach bo lamen va keo gan lam klnh dung djch SSC 4X10,05% Tween20 (pH 7,0) & nhi$t dOphong kinh, rLra lam antifade II, sau d~t lamen IEmtren BU'ae 10: quan sat tin hi$u huynh cua TB phoi tren kinh hit§n vi huynh quang KET QUA NGHIEN cuu vA BAN LU';N Sau tach chiet phan I~p dU'Q'cTB ph6i mau nqoai vi me va nhuorn TB b~n~ bQ kit Vysis MultiVysion PGT Multicolor probes, co probe gan d~e hieu trer NSTY Hinh 2: Hlnh anh nhuorn huynh quang mau TB phoi thai 56 555 (NST 56 13: ph6 mau do, NST 56 18: ph6 Aqua xanh nU'ac bit§n, NST 56 21: ph6 xanh 18 cay, NST Y: ph6 mau vang, NST X: ph6 mau xanh reu) 52 s6 6·2014 TAP CHi Y • DU'OC HOC QUAN SU' Hlnh anh tren cho th~y day la ph6i mang b¢ NST 46, XY cac NST 13, 18, 21 d~u c6 hieu, rnau vang c6 tin hieu cua NST Y va mot tin hieu rnau xanh reu cua NST X d~t ,,,.~~""", :h6 Them mien tren ~u huynh quar ii huynh quan; Hinh 3: Hinh anh nhu¢m huynh quang NST cua TB h6ng diu c6 nhan ph6i thai 56 556 Ket qua tren TB h6ng cau c6 nhan cua ph6i thai benh nhan nghien cuu 56 556 c6 J-JSTX va cac NST 13, 18, 21 d~u c6 tin hieu binh thLl'cYng.Trong tieu ban kh6ng t hien tin hieu rnau vang cua NST Y Qua phan tich ket qua cua 30 ph6i thai nqhien cuu, chunq t6i thu dLl'Q'cbang ket qua rotype cua NST 13, 18, 21 va X, Y nhu sau: SIT MABN CHAN E)OANADN PHOI TI,1DO TREN GEN SRY CHANE)oAN FISH K~TQuA SIEUAM 269 + 46,XY + 270 46,XX 271 46,XX 272 275 276 276 277 46, XX & + 46,XY + 46, XX J 21: ph6 xanh + 46, XY + y ' _10 J 278 280 + _j_ 46 XY -L- 46, XX KhOng xuat hien tin hi~u + NG + l -+ + f -i GHICHO ' - + '- _.j i~ 53 TAP CHI Y - DU'OC HOC QUAN SU' s6 6-2014 46,XX 11 295 296 + 46,XY + 12 297 + 46,XY + 13 298 + 46,XY + 14 15 300 301 + 46,XY + 16 303 NG + 17 + 307 46,XY + 18 308 46,XY + 19 20 309 21 310 22 311 23 312 46,XX 24 313 46,XX 314 + 46,XY + 25 315 + 46,XX + 26 27 316 28 317 29 318 46,XX 30 319 46,XX 46,XX + KhOngxu~t hien tin hi~u 46,XX 46,XX 46,XX 46,XY + 46,XY + 15 T8 hOng cau co nhan cua phoi thai mang NST XX va 13 h6ng cau co nhan cu: ph6i mang NST XV, Trong 30 ph6i nay, phoi kh6ng xuM hi$n tin hieu, Co th~ ph6i kh6ng pha va mang T8 va mang nhan, v$y cac probe kh6ng th~ gan vao NST Trong 15 phoi oU'Q'c ki§m chLrng bang ADN ph6i thai tl,l' mang NST dU'Q'c pnan I$p tLl' T8 phoi, chung t6i nhuQm c6ng cho 13/15 trU'cmg hop kl" nhuorn FISH cac phol dU'Q'c ki§m chLrng bang ADN ph6i thai tl,l' mang NST dU'Q'cnhuorn FISH d~u thay bQ NST xuat hlen NST X, di~u phu hop v&i k~t cua ADN phoi thai tl,i' va sleu am sau cac bat thU'ang NST va benn di truyer bang phuonq phap kh6ng can thlep Qua nghi€m CLrU nay, chLrng minh duoc mau ngo
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