PMC

Preparation of agar block smears. (a) Spore formations and other characteristic structures were checked by agar block examination of agar plates under a light microscope. (b) An agar block (15 by 15 mm) was cut using a sterile dissecting knife. (c) A coverslip (18 by 18 mm) was put onto the agar block after lactophenol cotton blue staining was performed. (d) The agar block with the coverslip was examined under a light microscope. (e) The block was dried in air until the thickness of the agar block reached 0.5 mm. (f) The agar block under the coverslip was filled with mounting medium.

Pour melted agar into a Petri dish (A) and embed tissues in agar (B). Cut out agar block (C) for slicing.

Conceptual design and operation. a Objective of proposed device showing the relative sizing of the 96-well plate and agar block. b Spheroids (red) grown in a 96-well plate (light grey) with liquid medium (blue). c Funnel manifold (green) with attached agar block (dark grey) mounted on a 96-well plate. d Centrifuge process using centripetal acceleration to move spheroids into the agar block. e Populated agar block removed from funnel manifold. f Agar block encapsulated and paraffin embedded after backfilling with liquid agar (light green). g Top and side view of a histological section of the agar block. Note: Not to scale, spheroids enlarged for clarity

The Song technique—agar stabilization bodies for a one step fully melting procedure. (A) Boiling agar 2% like in the molecular biologic laboratory for gels or in the kitchen. (B) The liquid agar is poured into the lid of a pipette box (waste material). (C) The solidified agar, which can be simply released from the mold is cut into plates of desired dimension by a scalpel. (D) Pouring the holes into an agar plate: This top pin tissue arrayer is placed into liquid agar. (E) After solidification of the agar the pins are withdrawn from the agar by turning the screws (D). (F) The agar plates (with or without preformed holes) are paraffinized in a standard automatic tissue processor. (G) The agar plate (agar stabilization body) is poured into a paraffin block. (H) Before drilling the holes into such a paraffin block and/or before filling the holes of a cast agar plate (D,E) sectioning of the block is recommended until the agar plate is in contact with the block surface. (I) The holes of the stabilization body can also be punched, e.g., with a manual tissue arrayer. (J) Agar stabilization body cast into a paraffin block with the holes being filled with PTCBs. (K) Agar plates can also be used and filled with PTCBs as a stand alone and may be cast into a paraffin block after melting. Note, this agar plate (asterisk) is thin (see insert) and gives stabilization only for one PTCB per hole. The advantage of this thin plate is the better release of air bubbles at melting. (L) A thick agar plate nearly completely surrounds the PTCBs at the entire length (see also insert (upper right corner) with a thick stabilization body (asterisk)). The holes can be filled with more than one PTCB to ensure an equal length and to prevent the thinning of the PTCBs in deeper sections. PTCBs of different diameters can be installed into the holes (inserts with arrowheads). The holes of the agar plate can be also constructed by punching manually with some more or less precise arrangement of the cores (arrow). Different diameters of the holes are possible. (M) Gap (arrows) between a PTCB and the surrounding agar stabilization body (a) before melting. (N) Gap (arrows) filled with paraffin after melting (a agar stabilization body). (O) Quick Ray manual tissue microarrayer set (Unitma, Seoul, South Korea). (P) Agar stabilization bodies of 1 mm, 1.5 mm, 3 mm, and 5 mm (Unitma, Seoul, South Korea) (Q) Fully automated tissue arrayer of Unitma (Seoul, South Korea) constructing PTMAs with preformed stabilization bodies.