TATA CARA NEKROPSI TIKUS dan PENGUJIAN PATOLOGI

Prof. Bambang Pontjo Priosoeryanto, Drh, MS, Ph D, APVet, Dipl. ACCM.Kepala Divisi Patologi

Departemen Klinik, Reproduksi & Patologi (KRP)Fakultas Kedokteran Hewan; Institut Pertanian Bogor

Disampaikan dalam acara Workshop Handling & Etika Penggunaan Hewan Coba serta PenandatangananMoU Program Studi S1 Farmasi Universitas Gunadarma, Rabu, 20 Februari 2019, Kampus Graha Simatupang,Universitas Gunadarma

Profil Prof. Bambang Pontjo Priosoeryanto, Drh, MS, Ph D, APVet, Dipl. ACCM.

Tempat/Tanggal lahir : Bandung, 28 Februari 1960NIP : 19600228 198601 1 001Pangkat : Pembina Utama Madya/ Guru BesarGolongan : IV/DJabatan : Kepala Divisi Patologi VeterinerFakultas : Kedokteran HewanDepartemen : Klinik, Reproduksi & Patologi (KRP)Divisi : Patologi VeterinerAlamat Kantor : Jalan Agatis, Kampus IPB Darmaga, BOGORTelepon dan Faksimili : (0251) 8421807 – kantor

E-mail : bpontjo4@gmail.com

Strata Tahun Bidang TempatS-1 (Drs. Med. Vet.) 1983 Kedokteran Hewan Institut Pertanian BogorDokter Hewan (Drh) 1984 Kedokteran Hewan Institut Pertanian BogorS-2 (MS) 1990 Sains Veteriner Institut Pertanian BogorS-3 (Ph.D) 1994 Pathology & Preventive Veterinary

MedicineUnited Graduate School of Veterinary Sciences, Yamaguchi University, Japan

Post Doktoral 1999 Neuropathology of Prion Diseases Institute for Neuropathology, Faculty of Medicine, Georg-August University, Goettingen, Germany

Brevet 2009 Ahli Patologi Veteriner (APVet.) Asosiasi Patologi Veteriner Indonesia (APVI)

2013 Diplomate Asian College Conservation Medicine ( DACCM)

Asian College Conservation Medicine, Japan

Keahlian Patologi Veteriner

Pemeriksaan Keadaan Luar

Periksa: 1. Keadaan Umum; 2. Jenis kelamin; 3. Kulit dan bulu; 4. Lubang-lubang kumlah; 5. Glandula mamari

Pemeriksaan Glandula Mamaria

• Tempatkan hewan dengan punggung menempel pada styrofoam

• Fiksir tiap kakinya dengan jarum pentul.

• Basahi seluruh kulit dan bulu bagian abdomen dan medial kaki dengan alkohol

• Buat sayatan di kulit sepanjang linea alba, mulai dari ujung dagu (regio mentalis) hingga ke tepi anterior tulang pelvis (pecten ossis pubis).

Tahap Nekropsi

Pembukaan kulit

• Kulit dipreparir hingga dapat dikuakkan ke samping tubuh, termasuk kulit di bagian atas dari kaki-kaki.

• Fiksir kulit dengan jarum pentul

• Sambil membuka kulit, dilakukan pengamatan pada subcutan

Penyayatan Kulit

Lokasi Sistem Limfatik

Tikus Betina Dengan Glandula Clitoris (GC: Gl. Clitoridis)

• Otot perut (dinding abdomen) digunting di linea alba, dimulai dari ujung tulang dada (processus xiphoideus) hingga pecten ossis pubis.

• Gunting otot perut dibawah kurvatura tulang rusuk dan di daerah sekitar paha, hingga otot-otot perut dapat dikuakkan ke kanan dan ke kiri

• Untuk lebih memudahkan mengamati organ-organ rongga perut, otot-otot perut disingkirkan.

Pembukaan Ruang Abdomen

Pembukaan ruang abdomen dan pemeriksaan umum

Pemeriksaan Ruang Abdomen

Rongga Perut dengan Berbagai Organ

Sistem Pencernaan dan Limpa

Pemotongan oesophagus (OE) agar dapat mengeluarkan organ-organ pencernaan. CX: Cartilago xiphoidea; Spleen

Sistem Pencernaan dan Limpa

Lambung Tikus dan Bagian-bagiannya

Ginjal dan Kelenjar Adrenal

Penis dikeluarkan bersama dengan kelenjar asesorius (gl. vecicularis, gl. vesicula seminalis, gl. prostate, gl. bulbourethralis), vesica urinaria, urethra

dan penis. DD: Ductus deferens.

Urogenitalia Tikus Jantan

Testis, Epididimis dan Ductus deferens

Urogenitalia Tikus Jantan

Kelenjar asesorius pada tikus jantan. P: gl. Prostate, GC: gl. coagulationis, GV: gl. Vecicularis, GB: Gg. Bulbourethralis,

U: Urinary bladder.

Urogenitalia Tikus Jantan

Organ-organ di ruang abdomen dan pelvis tikus betina. R: Rectum, OV: Ovarium, SP: Symphysis pelvina.

Alat Reproduksi Tikus Betina

1.Tulang rusuk terakhir dipotong ke depan menuju arkus tulang sternum

2.Pemotongan dilakukan pada sisi kanan maupun kiri, kemudian perlekatan dengan difragma dipreparir, sehingga tulang sternum dan sebagian tulang rusuk berbentuk segitiga dapat dibuang

Pembukaan Rongga Dada

CX: Cartilago xiphoidea. DI: Diaphragma.

Pembukaan Rongga Dada

Pemeriksaan Timus

Pengeluaran organ-organ rongga dada dalam satu rangkaian: lidah, esophagus dan trakhea

Pengeluaran Organ-organ Rongga Dada

•Buat sayatan pada kulit kepala tepat dibagian tengah dan berakhir sejajar telinga

•Buang kulit dan otot-otot di bagian dorsal dan kaudal kranium

Pembukaan Kranium

Pembukaan Kraniumdan Pengambilan Otak

Pembukaan Kranium

Pemeriksaan gl. Harderian

Pemeriksaan hyphophysa setelah otak

diambil

Pemeriksaan kelenjar ludah dan limfonodus. LNN: Lymph nodes. GSL: gl. sublingualis. GSM: Gl. submaxillaris.

Pemeriksaan Kelenjar Ludah

GLE: Gl. lacrimalis extraorbitalis. GP: Gl. parotis

Pengambilan Mata

GT: gl. Thyreoidea, L: Larynx, T: Trachea, M: Musculus masseter.

Bagian Ventral Leher

Nervus ischiadicus (NI) diantara otot paha medial

Sampling Organ

4 Submitting Multiple Sites

How To and How Not To Submit your Biopsy Specimens DA Kamstock, DVM, DACVP, LL Debuse, EJ Ehrhart, DVM, DACVP, BE Powers, DVM, DACVP

Colorado State University Veterinary Diagnostic Laboratory

3 Packaging

Routine tissue fixation = 1:10 tissue to neutral buffered formalin

For appropriate fixation, 0.5 – 1.0 cm tissue thickness is

recommended

Bread loafing (incomplete parallel cuts at a minimum of 2cm

apart) can be performed on large specimens (be sure to avoid

complete transection or too many cuts which can both result in

loss of tissue orientation!)

Specimens can be held to fix (at least 24 hours) at your clinic

prior to sending to the lab to avoid shipping large volumes of

formalin which can be expensive and increase the risk of leaking

2 Tissue Fixation

6 Denoting Margins

7 Other Things to Avoid

8 What Else Should I know?

It is important for you to realize that after all is said and done

the pathologist typically evaluates 1 to 4, 5um thick sections

from the entire specimen which is submitted (A).

Our staff and pathologists are here to assist you. If you have

questions about how best to submit your sample or have

questions regarding any other issues, please contact the lab -

(970)-297-1281.

Additional information on the CSU-VDL can be found on the

web at www.dlab.colostate.edu Visit Us!

1 Clinical Information

Please provide signalment and pertinent clinical information

Certain lesions occur more commonly in different species and

certain breeds

Anatomical location of a lesion, as well as clinical appearance and

progression, may also be critical information to allow your

pathologist to provide you with the best possible diagnosis and/or

differentials

If you have a specific question, are concerned about a possible

disease process, or have a list of differentials you’d like to rule out,

please indicate such.

Again, please make every effort to provide this necessary

information in the designated areas on the CSU-VDL biopsy

submission form. It will help us help you help your patients.

5 Endoscopic Biopsies

B. Large samples can be held

and fixed at your clinic prior to

submitting to the lab to help

avoid shipping large volumes of

formalin which may be costly and

hazardous

B

C. This is an example of an ~20cm

diameter mass lesion which was fixed

at the clinic and subsequently sent to

the lab in a plastic, labeled, zip lock

bag devoid of any formalin. (bar =

~2.5 cm)

C

A. Incomplete parallel cuts at a

minimum of 2cm apart (bread

loafing) can be utilized to assist

with appropriate tissue fixation

for large specimens

A

Surgical Ink 1. Ink the area of interest

2. Be sure to ink prior to bread loafing (if

needed) 3. Allow ink to

begin drying before placing the specimen in

formalinTagging

1. Used to indicate margins or for orientation

2. Use variable numbers and/or colors of suture

3. Provide a clear description on the

submission form denoting what the sutures

indicate (i.e. one suture = cranial margin)

Tumor Bed Samples

1. Submission of samples from the post-surgical

bed 2. Any tumor in these

specimens is evidence of remaining microscopic

disease 3. Similar to

“submitting multiple sites” clearly label and submit

each region individually

Formalin filled jars containing specimens should be placed in

a plastic bag, box, or other container with absorbent material to

absorb any leakage (A)

Paperwork should be placed in a separate plastic bag to avoid

contact with formalin if leaking does occur. Such contact can

result in altered and illegible paperwork (B)

Your fresh sample size should never be larger than the most

narrow portion of the jar in which you are submitting it (C). If it

is, this will require cutting plastic jars or breaking glass jars

(undesirable) in order to retrieve the tissue which may become

altered in the process.

YES

The optimal method by which to submit

multiple lesions from a single animal is to

submit each specimen individually in its own

respective and appropriately labeled jar. This

should be reflected on the submitted

paperwork.

If multiple specimens are submitted in a

single container (which is less ideal) there

needs to be some method of tissue

identification (i.e. suture) to denote

specimens relative to respective anatomic

site. YES

suture

NO NO YES

Do not submit endoscopic

biopsies wrapped in gauze

sponges. Specimens may

become lost or may be crushed

during the attempted retrieval

process. It is better to submit

the specimen free floating in the

jar than with gauze or any other

material

The optimal method by which to

submit an endoscopic biopsy is to

place it in a screen cassette after

which the cassette should be placed in

an appropriately labeled formalin filled

jar. If individual cassettes are labeled

properly (sharpie or no2 pencil),

multiple cassettes can be place in one

jar.

Do not place endoscopic

biopsies on fragments of

cardboard. Specimens will

either float off or, if adhered,

tissue will slough off during

retrieval and/or may be

associated with cardboard

fibers

A. From

mass to

block to

slide

Our staff

is here

working

hard for

you!

Please help keep our technicians

fingers safe and DO NOT submit

specimens with needles for any

reason!

NO

NOPlease do no staple, suture, or pin

tissue to cardboard. It can damage

tissue and prevent appropriate margin

assessment

YES NO

YES

NO

A

CB

Sampling Organ• Mengumpulkan sampel se-aseptik mungkin

• jaringan

• aspirat

• kultur darah

– Alat nekropsi steril• Disinfeksi cold pack

• Alat autoclave

• Alkohol and pembakar

• Masukkan sampel yang representatif

• Ambil sampel dari yang rusak

• Ambil sample dari yang segar dan baik

• Fiksatif Buffered Neutral Formalin 10%

• Volume Fiksatif : Organ = 10 : 1

• Waktu fiksatif yang cukup minimal 48 jam sebelum prosesing jaringan

Sampling Organ

Jaringan• Koleksi sebesar genggaman (jika kurang, jaga kelembaban)

• Hanya jaringan yang dikirim untuk histopatologi yang harus diberi formalin

• Jaringan dapat dikirim dalam:– Whirl Pac– Mangkok fecal/ urine yang bersih (belum digunakan) atau

kontainer plastik.– Snap cap tube– Red top tube

• Jaringan harus tetap lembab– (cukup untuk melindungi jaringan)– Bungkus dalam steril 4X4 soaked dalam salin steril

JARINGAN

Contoh PengirimanSample yang Buruk

Contoh Pengiriman Sample yang Baik

PENGUJIAN PATOLOGI

MAKROSKOPIS MIKROSKOPIS

Patologi Anatomi Histopatologi

Sitopatologi

Histokimia (Pewarnaan Khusus)

Imunohistokimia (Reaksi Ag-Ab)

Elektron Mikroskopi (SEM & TEM)

ANALISI YANG DIPAKAI 1. Deskriptif2. Skoring Lesio dengan metode Histoskor (Intensitas x

Distribusi), yang dilanjutkan dengan Analisis Statistika

Studi Infeksi Virus Parvovirus - MyokarditisIntranuclear inclusion body

Sarang radang

AnalisisHistopatologi

TahapanKerusakan

otot jantungakibat

Kardiotoksin

Studi HistopatologiKinetika Tumor

Basal Sel Tumor

TC

HP

StudiTransplantasiSel Tumor

Pada Nude Mouse

Studi Tumor Imunologi

ImunohistokimiaAntibodi-Antikeratin

• Signalemen

• Anamnese

• Temuan Patologi Anatomi

• Histopatologi

• Diagnosa/ Diferensial diagnosa

Penulisan Laporan

REFERENSI

• Henrik Elvang JensenDepartment of Veterinary Pathobiology, Royal Veterinary and Agricultural University, Copenhagen, Denmark

• Vincenzo Covelli - Guide to the Necropsy of the Mouse

• Dan lain lain

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