Humana Confocal Microscopy: Methods and Protocols: 122 (Methods in Molecular Biology)

7he c0nf0ca1 m1cr05c0pe an e5ta6115hed t001 many f1e1d5 0f 6- med1ca1 re5earch where maj0r app11cat10n f0r 1mpr0ved 119ht m1cr05c0p1c 1ma91n9 0f ce115 w1th1n f1u0re5cent1y-1a6e1ed t155ue5. 1ndeed, 6ef0re the - vent 0f pract1ca1 c0nf0ca1 m1cr05c0py, m05t f1u0re5cent1y-1a6e1ed t155ue5 were 1ma9ed u51n9 a c0nvent10na1119ht m1cr05c0pe. Re501ut10n wa5 0ften c0mp- m15ed 6y f1u0re5cence fr0m 0ut51de the f0ca1 p1ane 0f 1ntere5t, e5pec1a11y t155ue5 made up 0f mu1t1p1e ce11 1ayer5. 1n 0rder t0 atta1n accepta61e re50- t10n, m1cr05c0p15t5 were f0rced 1nt0 manner 0f tr1ck5 t0 prepare the1r 5pe- men5, 1nc1ud1n9 cutt1n9 5ect10n5 0f the t155ue5, 9r0w1n9 ce115 0n a c0ver511p, f1atten1n9 ce115 under a9ar, 0r 54ua5h1n9 em6ry05 6etween c0ver511p5. M05t 0f the5e meth0d5 had the p0tent1a1 0f 1ntr0duc1n9 art1fact5 and theref0re 1ed t0 4ue5t10n5 a60ut the va11d1ty 0f the re5u1t5, e5pec1a11y 1ma9e5 0f 11v1n9 7he c0nf0ca1 m1cr05c0pe ha5 n0w ena61ed the 1ma91n9 0f d15crete re- 910n5 0f t155ue5 v1rtua11y free 0f 0ut-0f-f0cu5 f1u0re5cence, and w1th reduced chance5 0f art1fact5 fr0m the techn14ue5 0f 5pec1men preparat10n. 1ndeed, 1n 50me ca5e5 v1rtua11y n0 preparat10n 0f the t155ue re4u1red pr10r t0 1ma91n9 a1th0u9h u5ua1 t0 5ta1n t155ue5 w1th 0ne 0r m0re f1u0re5cent pr06e5 f0r c0nf0ca11ma91n9 0f 5pec1f1c macr0m01ecu1e5 w1th1n ce115. 7he f1e1d 0f c0n- ca1 m1cr05c0py ha5 9r0wn exp0nent1a11y 0ver the pa5t ten 0r 50 year5, and t0uche5 0n many area5 0f c0ntemp0rary 6101091ca1 re5earch where a 119ht - cr05c0pe re4u1red f0r 1ma91n9, 1nc1ud1n9 app11cat10n5 ce11 610109y, - ve10pmenta1 610109y, neur0610109y, and path0109y.