IMUNISASI Usaha pencegahan suatu penyakit dengan pemberian vaksin (imunitas aktif) atau antisera (imunitas pasif).

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• ANTISERA • Serum yang telah mengandung antibodi • Contoh : • ATS (Anti Tetanus Serum) • ABU (Anti Bisa Ular)

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• Vaksinasi • Suatu imunoterapi anti-mikrobial, melibatkan pengaktifan sistem imun untuk merespon agen penyebab infeksi. • Apakah perlu imunisasi pada bayi? • Bayi yang baru lahir sudah mendapat antibodi dari ibunya usia 6 bulan
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• Setelah usia 6 bulan. bayi memerlukan imunisasi • Tetapi tidak harus diimunisasi • Persetujuan orang tua • Memberi penjelasan vaksin yang akan diberikan 4 .

Vaksin adalah preparat antigenik yang digunakan untuk meningkatkan imunitas terhadap suatu penyakit. Proses distribusi dan pemberiannya disebut vaksinasi. Pasteur menemukan vaksin terhadap smallpox. Istilah ini untuk memberi penghargaan pada Edward Jenner yang meneliti tentang cowpox. vacca=sapi. Istilah vaksin berasal dari bahasa latin. 5 .

• Vaksin dapat untuk profilaktik (untuk pencegahan atau memperbaiki efek dari infeksi oleh patogen) atau terapeutik (untuk pengobatan. misal terhadap kanker). 6 .

yang dilemahkan. 7 . • Atau dapat berupa organisme yang dimatikan atau diinaktifkan atau produk murni yang berasal dari bagian atau fragmen organisme.• Tipe vaksin • Vaksin dapat berupa virus atau bakteri hidup.

• Bahan kimia yang digunakan : formalin atau fenol • Vaksin tipe ini menimbulkan respon imun yang tidak komplit atau yang pendek waktunya dan memerlukan penggunaan secara boster. bubonic plague dan hepatitis A. • Contoh : vaksin flu. 8 . Inaktivasi : dibuat dari mikroorganisme virulen yang dimatikan dengan bahan kimia atau pemanasan. kolera.• Ada 4 tipe vaksin tradisional : • 1.

• 2. Mikroorganisme hidup yang dikurangi virulensinya. adalah : mikroorganisme hidup yang dikultivasi dibawah kondisi yang tidak enak bagi sifat virulennya. Memproduksi urease nyeri. Misal : virus polio dikultur (ditumbuhkan dalam telur ayam yang sudah dibuahi) Helicobacter pylori. measles. • Contoh : yellow fever. fimbrie nya akan hilang bakteri gundul • Vaksin tipe ini menimbulkan respon imun lebih panjang dan lebih banyak digunakan untuk orang dewasa sehat. Bakteri ini dikultur berkali-kali (20-50 X). dan mumps 9 . hidup di lambung. rubella.

berupa fragmen dari mikroorganisme. tipe ini dibuat dari senyawa toksik dari mikroorganisme yang diinaktifkan. dipteri • 4.• 3. • Senyawa tersebut pada keadaan tertentu lebih dapat menimbulkan penyakit dibanding dengan mikroorganisme itu sendiri. Sub unit. dibuat dari protein permukaan virus 10 . • Contoh : tetanus. • Misal : vili • Contoh : vaksin HBV. yang dapat menimbulkan respon imun. Toksoid.

VIRION 11 .

Eukaryote versus prokaryote 12 .

tetapi dari strain BCG. 13 .• Vaksin tuberkulosis hidup bukan merupakan strain TB yang dapat menyebabkan penyakit menular. • Di USA. vaksin ini jarang digunakan.

maka sistem imun dapat mengenali polisakarida tersebut sebagai antigen protein • Contoh : Vaksin Haemophilus influenzae tipe B 14 .• Beberapa vaksin inovatif (modern) yang sedang dikembangkan dan juga dalam penggunaan : • 1. Konjugat . Dengan melekatkan polisakarida pada protein (misal toksin).bakteri tertentu mempunyai • lapisan luar berupa polisakarida yang sifat imunogeniknya lemah.

dengan mengkombinasi sifat fisiologi satu mikroorganisme dengan DNA dari mikroorganisme yang lain. • Imunitas dapat dibentuk terhadap penyakit yang mempunyai proses infeksi yang komplek 15 .• 2. Vektor rekombinan.

Akhir-akhir ini. • Transformasi DNA bakteri atau virus kedalam sel manusia atau binatang. tipe baru dari vaksin dikembangkan dari DNA agen infeksi. Vaksin DNA.• 3. 16 .

jika ada patogen yang biasanya mengekspresikan protein tersebut masuk kedalam hospes.• Karena sel tersebut dapat hidup lama. mereka akan diserang dengan segera oleh sistem imun. 17 . • Salah satu keuntungan vaksin DNA adalah sangat mudah diproduksi dan disimpan.

• 4. Terdiri dari 10-15 asam amino. yang digunakan sebagai vaksin. Vaksin sub unit • Yang bisa menimbulkan respon imun. 18 . bagian yang muncul di permukaan (epitop). Vaksin sintetik • 5.

rotavirus.• Di USA. meningitis dan pneumonia. rubella. 19 . diptheria. memberikan rekomendasi vaksinasi rutin pada anakanak terhadap hepatitis A. influenza. the Advisory Committee on Immunization Practices. mumps. tetanus. pertussis. chicken pox. measles. polio. hepatitis B.

dengan tujuan untuk pencegahan terhadap bermacam-macam penyakit.• Banyaknya jumlah vaksin dan boster yang direkomendasikan (sampai dengan 24 injeksi pada umur 2 tahun) telah menjadi masalah untuk pencapaian yang penuh. 20 . Vaksin kombinasi telah dipasarkan (misal vaksin Prevnar dan ProQuad).

• Vaksin yang direkomendasikan untuk pemberian ulang seumur hidup : measles. dan pneumonia. 21 . influenza. Seringkali wanita hamil diskrining untuk terus resisten terhadap rubella. tetanus.

yang lebih mematikan untuk golongan tersebut. 22 .• Vaksin yang direkomendasikan untuk usia tua : pneumonia dan influenza.

(pengawet) yang dimetabolisme menjadi etilmerkuri. Contoh : vaksin influenza. penggunaan thimerosal sudah banyak berkurang. DTP (diphtheria.• Kontroversi Vaksin • Sejumlah vaksin. • Sejak tahun 1997. tetanus dan pertussis). mengandung thimerosal. • Di USA telah ditetapkan hukum yang melarang penggunaan thimerosal dalam vaksin untuk anakanak. 23 .

menimbulkan efek samping autisme.• Penelitian tentang thimerosal : • Laporan akhir tahun 1990. Mumps. Rubella) dan hepatitis B. 24 . • Laporan pada tahun 2004-2005. vaksin MMR (Measles. banyaknya penderita mumps (gondok) pada anak-anak dan orang tua.

yang diserang sel Tnya • 2. sehingga sel T terganggu. Penerima transplantasi.• Efektifitas yang rendah dari pemberian vaksin • 1. seperti AIDS. Pada hospes yang menderita kelainan genetik • Tidak mempunyai ADE (Adenin Deaminase). menderita seperti AIDS atau lebih dikenal SCID (Severe Complex Immuno Deficiency) 25 • 4. juga propanolol • 3. Pada hospes yang tak mampu membentuk antibodi. sering diberi obat . misal : golongan obat steroid. Pada hospes yang menerima obat yang dapat menurunkan sistem imun.

hasilnya lebih baik daripada penyakit lainnya) • 2. Penyakitnya (untuk beberapa vaksinasi penyakit tertentu. Skedul waktu pemberian • 4. tergantung pada beberapa faktor : • 1. Etnik atau genetik 26 .• Efektifitas atau penampilan vaksin. Strain dari vaksin • 3. Individual • 5.

orang tersebut terkena penyakit. masih ada kemungkinan. • 2. Sistem imun hospes. Sistem imun hospes tidak mempunyai sel B • Contoh : Polio 90-100% Tetanus >90% Diphteria 87-96% 27 . Adanya imunitas yang direndahkan (diabetes. penggunaan steroid.• Efikasi Vaksin • Vaksin tidak menjamin perlindungan yang komplet. infeksi HIV) • 3. • Bisa disebabkan : • 1.

Beberapa germ dapat bermutasi (virus penyebab demam dan influenza). 28 . sistem imun manusia tidak sempurna. dan dalam beberapa kasus sistem imun masih belum mampu untuk memerangi infeksi.• Bahkan apabila hospes mengembangkan antibodi.

Hepatitis B • 4. karena bahan pengawetnya 29 .• Vaksin yang dapat meningkatkan resiko terjadinya diabetes melitus • 1. karena kelainan genetik • Vaksin . Hemophilus • 3. Pertussis • 2. Tuberculosis (BCG) • Bisa berasal dari hospes atau dari vaksinnya • Hospes.

Transverse myelitis and multiple sclerosis diketahui ada hubungannya dengan vaksin. 30 . seperti Acute disseminated encephalomyelitis. Guillain-Barré syndrome.• Efek Samping • Beberapa penyakit autoimun.

Contoh : Mycobacterium leprae Hanya bisa hidup di hewan trenggiling.• • • • • • • • • Produksi vaksin Melalui beberapa tahap 1. Kultivasi secara in vivo dan invitro Tak semua agen infeksi bisa dikultivasi in vitro. jika di luar tubuh manusia Virus MMR bisa dikultivasi in vivo di dalam embrio dengan kultur jaringan atau sel Vibrio cholerae dapat dikultivasi in vitro31 .

terhadap temperatur.• Pemanenan dan pemurnian Bisa dilakukan secara sentrifugasi dan kromatografi • Formulasi Untuk menjaga konsistensi vaksin. agar kalau didinginkan tidak membeku 32 . maka ditambahkan gliserin.

tidak pirogenik.00125% • Elektrolit dan bufer. tentunya dapat meningkatkan imunogenik suatu antigen 33 . untuk menjaga pH larutan vaksin • Adjuvan. inert. tetapi bisa menimbulkan resistensi Thimerosal. suatu material yang dapat meningkatkan respon imun Syarat : tidak imunogenik. mudah didegradasi. digunakan dalam jumlah sangat kecil. sekitar 0.• Pengawet • Bisa digunakan antibiotik.

hanya sekali pemberian • Mudah diberikan.• Vaksin yang ideal • Dapat digunakan untuk semua umur • Proteksi seumur hidup. karena injeksi bisa menimbulkan trauma pada anak-anak. • Tidak ada efek samping • Stabil • Tersedia dalam jumlah besar • Murah 34 . jika mungkin per oral.

Liposomes and Immunostimulating complexes (ISCOMS) 35 .• • • • • • Adjuvan Senyawa peningkat respon imun. Contoh : Adjuvants in common use: 1. Garam Aluminium 2.

oil and water • Too toxic for man • Induces a good cell mediated immune response. Incomplete Freund's adjuvant as above. Muramyl di-peptide • Derived from Mycobacterial cell wall. 36 . IL-12 and Interferon-gamma. Cytokines • IL-2.• 3. Complete Freunds adjuvant is an emulsion of Mycobacteria. • 5. but without Mycobacteria. • 4. • 6.

• Imunoterapi yang lain • Kanker • Imunoterapi BCG 37 .

jerawat. • Imunoterapi injeksi menggunakan antigen mumps.• Imunoterapi topikal menggunakan krim (imiquimod) untuk kutil. candida atau trichophytin untuk mengobati kutil. squamous cell cancer. cutaneous lymphoma. 38 . basal cell cancer. dan superficial malignant melanoma.

dan sel dendritik. 39 . sel TCD8+. diambil dari pasien.• Imunoterapi dengan sel dendritik • Tujuannya untuk mengaktifkan respon sitotoksik terhadap suatu antigen. juga sel B). dikembalikan lagi ke pasien. dicampur dengan antigen ataupun dengan vektor virus. Sel dendritik yang sudah aktif. sel tersebut akan menyajikan antigen ke sel limfosit efektor (sel TCD4+. • Sel dendritik (sel APC).

• Imunoterapi adoptif menggunakan sel T • Terapi ini menggunakan respon sitotoksik oleh sel T untuk menyerang kanker. 40 . kemudian dipindahkan lagi ke pasien tersebut. Sel T dari pasien yang mempunyai reaktivitas secara alami maupun genetik pada pasien kanker. ditingkatkan aktivitasnya.

dan ditingkatkan aktivitasnya secara in vitro dengan IL-2. anti-CD3 dan kadar tinggi 41 .• Tumor pasien dipanen.

• Imunosupresi • Menekan respon imun abnormal dalam penyakit autoimun atau usaha untuk mengurangi respon imun normal untuk mencegah penolakan terhadap sel atau organ yang ditransplantasikan. 42 .

• Imunoterapi juga digunakan untuk mengobati alergi. • Obat alergi. • Imunoterapi tidak efektif untuk semua orang. 43 . antihistamin atau kortikosteroid hanya mengobati simtom penyakit alergi. dengan mereduksi sensitivitas terhadap alergen. sedangkan imunoterapi dapat memodifikasi secara alami penyebab penyakit alergi.

tetapi efikasinya masih kalah dibanding ASI 44 .• Perbedaan ASI dengan susu formula • ASI mengandung antibodi. susu formula tidak • Walau ada usaha. sapi memproduksi laktalbumin dengan rekayasa genetik. susu formula tidak • ASI mengandung laktalbumin.

The proteins are then transferred to a membrane (typically nitrocellulose or PVDF). 45 . immunoblot) is an analytical technique used to detect specific proteins in a given sample of tissue homogenate or extract. where they are probed (detected) using antibodies specific to the target protein.• Western blot • The western blot (alternatively. It uses gel electrophoresis to separate native or denatured proteins by the length of the polypeptide (denaturing conditions) or by the 3-D structure of the protein (native/ non-denaturing conditions).

immunogenetics and other molecular biology disciplines. 46 . Commercial antibodies can be expensive. biochemistry. although the unbound antibody can be reused between experiments. This method is used in the fields of molecular biology.• There are now many reagent companies that specialize in providing antibodies (both monoclonal and polyclonal antibodies) against many thousands of different proteins.

and resistance to solvents. it is used generally in applications requiring the highest purity. it has an easier melt process because of its relatively low melting point of around 177°C. Compared to other fluoropolymers. 47 .• Polyvinylidene Fluoride. • PVDF is a specialty plastic material in the fluoropolymer family. HYLAR or SYGEF. bases and heat and low smoke generation during a fire event. or PVDF is a highly non-reactive and pure thermoplastic fluoropolymer. strength. It is also known as KYNAR. acids.

It is available as piping products.78) and low cost compared to the other fluoropolymers. tubing. plate and an insulator for premium wire.• It has a relatively low density (1. as well as in lithium ion batteries. molded or welded and is commonly used in the chemical. sheet. 48 . It can be injected. semiconductor. films. medical and defense industries.

Detection of RNA is termed northern blotting. 49 . • The method originated from the laboratory of George Stark at Stanford. Neal Burnette and is a play on the name Southern blot. The name western blot was given to the technique by W.• Other related techniques include using antibodies to detect proteins in tissues and cells by immunostaining and enzyme-linked immunosorbent assay (ELISA). a technique for DNA detection developed earlier by Edwin Southern.

it should be noted that bacteria. 50 . virus or environmental samples can be the source of protein and thus Western blotting is not restricted to cellular studies only.• Steps in a Western blot • Tissue preparation • Samples may be taken from whole tissue or from cell culture. However. using a homogenizer (smaller volumes). solid tissues are first broken down mechanically using a blender (for larger sample volumes). In most cases. or by sonication. Cells may also be broken open by one of the above mechanical methods.

salts. and buffers may be employed to encourage lysis of cells and to solubilize proteins.• Assorted detergents. • A combination of biochemical and mechanical techniques – including various types of filtration and centrifugation – can be used to separate different cell compartments and organelles. 51 . Protease and phosphatase inhibitors are often added to prevent the digestion of the sample by its own enzymes.

The nature of the separation depends on the treatment of the sample and the nature of the gel. or a combination of these factors. molecular weight. electric charge. • The proteins of the sample are separated using gel electrophoresis. 52 .• Gel electrophoresis • Immunoblot (Western blot) analysis of proteins separated by SDS-PAGE gradientgel electrophoresis. Separation of proteins may be by isoelectric point (pI).

disulfide bonds [S-S] to sulfhydryl groups [SH and SH]) and thus allows separation of proteins by their 53 molecular weight.• By far the most common type of gel electrophoresis employs polyacrylamide gels and buffers loaded with sodium dodecyl sulfate (SDS). . SDS-PAGE (SDS polyacrylamide gel electrophoresis) maintains polypeptides in a denatured state once they have been treated with strong reducing agents to remove secondary and tertiary structure (e.g.

The lower the acrylamide concentration the better the resolution of higher molecular weight proteins.• Sampled proteins become covered in the negatively charged SDS and move to the positively charged electrode through the acrylamide mesh of the gel.the greater the acrylamide concentration the better the resolution of lower molecular weight proteins. The concentration of acrylamide determines the resolution of the gel . Smaller proteins migrate faster through this mesh and the proteins are thus separated according to size (usually measured in kilo Daltons. kDa). 54 . Proteins travel only in one dimension along the gel for most blots.

One lane is usually reserved for a marker or ladder. typically stained so as to form visible. coloured bands. a commercially available mixture of proteins having defined molecular weights. These different rates of advancement (different electrophoretic mobilities) separate into bands within each lane.• Samples are loaded into wells in the gel. proteins migrate into it at different speeds. 55 . When voltage is applied along the gel.

.• Samples are loaded into wells in the gel. When voltage is applied along the gel. a commercially available mixture of proteins having defined molecular weights. One lane is usually reserved for a marker or ladder. coloured bands. These different rates of advancement (different electrophoretic mobilities) separate into bands within 56 each lane. typically stained so as to form visible. proteins migrate into it at different speeds.

they are moved from within the gel onto a membrane made of nitrocellulose or polyvinylidene difluoride (PVDF). . bringing the 57 proteins with it. and a stack of tissue papers placed on top of that.• Transfer • In order to make the proteins accessible to antibody detection. The entire stack is placed in a buffer solution which moves up the paper by capillary action. The membrane is placed on top of the gel.

• Another method for transferring the proteins is called electroblotting and uses an electric current to pull proteins from the gel into the PVDF or nitrocellulose membrane. The proteins move from within the gel onto the membrane while maintaining the organization they had within the gel. As a result of this "blotting" process, the proteins are exposed on a thin surface layer for detection (see below).

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• Both varieties of membrane are chosen for their non-specific protein binding properties (i.e. binds all proteins equally well). Protein binding is based upon hydrophobic interactions, as well as charged interactions between the membrane and protein. Nitrocellulose membranes are cheaper than PVDF, but are far more fragile and do not stand up well to repeated probings.
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• The uniformity and overall effectiveness of transfer of protein from the gel to the membrane can be checked by staining the membrane with Coomassie or Ponceau S dyes. Coomassie is the more sensitive of the two, although Ponceau S's water solubility makes it easier to subsequently destain and probe the 60 membrane as described below.

and both antibodies and the target are proteins. with a minute percentage of detergent such as Tween 20. steps must be taken to prevent interactions between the membrane and the antibody used for detection of the target protein. Blocking of non-specific binding is achieved by placing the membrane in a dilute solution of protein typically Bovine serum albumin (BSA) or nonfat dry milk (both are inexpensive).• Blocking • Since the membrane has been chosen for its ability to bind protein. 61 .

there is no room on the membrane for it to attach other than on the binding sites of the specific target protein. Thus. 62 . when the antibody is added. and eliminates false positives. This reduces "noise" in the final product of the Western blot.• The protein in the dilute solution attaches to the membrane in all places where the target proteins have not attached. leading to clearer results.

For a variety of reasons. although there are now one-step detection methods available for certain applications. 63 . this traditionally takes place in a two-step process. which when exposed to an appropriate substrate drives a colourimetric reaction and produces a colour.• Detection • During the detection process the membrane is "probed" for the protein of interest with a modified antibody which is linked to a reporter enzyme.

with warmer temperatures being associated with more binding. a dilute solution of primary antibody (generally between 0. 64 . both specific (to the target protein. Typically. the solution is composed of buffered saline solution with a small percentage of detergent. and sometimes with powdered milk or BSA. The antibody solution and the membrane can be sealed and incubated together for anywhere from 30 minutes to overnight.5 and 5 micrograms/ml) is incubated with the membrane under gentle agitation.• After blocking. the "signal") and non-specific ("noise"). It can also be incubated at different temperatures.

this is part of the immune response. Normally.• Two step • Primary antibody • Antibodies are generated when a host species or immune cell culture is exposed to the protein of interest (or a part thereof). whereas here they are harvested and used as sensitive and specific detection tools that bind the protein directly. 65 .

an anti-mouse secondary will bind to just about any mouse-sourced primary antibody." etc. tends to be referred to as "anti-mouse." "anti-goat. Antibodies come from animal sources (or animal sourced hybridoma cultures). 66 . directed at a species-specific portion of the primary antibody. and due to its targeting properties.• Secondary antibody • After rinsing the membrane to remove unbound primary antibody. This is known as a secondary antibody. the membrane is exposed to another antibody.

and provides far more consistent results. This means that several secondary antibodies will bind to one primary antibody and enhance the signal. The secondary antibody is usually linked to biotin or to a reporter enzyme such as alkaline phosphatase or horseradish peroxidase.• This allows some cost savings by allowing an entire lab to share a single source of mass-produced antibody. 67 .

• Most commonly. A sensitive sheet of photographic film is placed against the membrane. and exposure to the light from the reaction creates an image of the antibodies bound to the blot. 68 . a horseradish peroxidase-linked secondary is used in conjunction with a chemiluminescent agent. and the reaction product produces luminescence in proportion to the amount of protein.

quicker and cheaper this method is now rarely used. such as labeling an antibody-binding protein like Staphylococcus Protein A with a radioactive isotope of iodine. • A third alternative is to use a radioactive label rather than an enzyme coupled to the secondary antibody.• As with the ELISPOT and ELISA procedures. 69 . the enzyme can be provided with a substrate molecule that will be converted by the enzyme to a colored reaction product that will be visible on the membrane (see the figure below with blue bands). Since other methods are safer.