FIKOSIANIN Spirulina
SIMPULAN DAN SARAN Simpulan
Simpulan
1. Spirulina platensis dapat dikultur dengan menggunakan nutrien ekonomis murah (modifikasi nutrien teknis MT) dengan intensitas cahaya sebesar 3000 lux pada skala massal untuk memproduksi fikosianin sebagai bahan imunostimulan.
2. Fikosianin dari Spirulina platensis dapat digunakan sebagai imunostimulan dalam meningkatkan respons imun non spesifik,
performance pertumbuhan dan resistensi juvenil ikan kerapu bebek terhadap infeksi bakteri V. alginolyticus dengan pemberian fikosianin pada dosis 250 mg kg-1 pakan.
3. Pemberian fikosianin 250 mg kg-1 pakan selama 14 hari meningkatkan respons imun non spesifik dan resistensi juvenil ikan kerapu bebek terhadap infeksi bakteri V. alginolyticus.
Saran
1. Perlu dicari komposisi perbandingan nitrogen dan fosfor, serta penambahan NaHCO3 yang optimum dalam media kulur Spirulina
platensis untuk menghasilkan fikosianin yang lebih tinggi.
2. Untuk meningkatkan respons imun non spesifik dan pertumbuhan ikan kerapu bebek dapat diaplikasikan penambahan fikosianin sebesar 250 mg kg-1 pakan dan dapat diaplikasikan selama 14 hari berturut-turut.
3. Penelitian ini masih membuka peluang pengkajian terhadap efektifitas ekstrak kasar fikosianin tanpa tahap pemurnian lebih lanjut, ekstrak air panas dan dinding sel Spirulina terhadap peningkatan respons imun non spesifik pada ikan kerapu bebek dan resistensinya terhadap bakteri patogen dan melakukan pengamatan terhadap jumlah penurunan bakteri patogen sebelum dan setelah infeksi untuk melihat korelasinya secara langsung.
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93 Lampiran 1. Prosedur kerja pengukuran beberapa parameter
1. Pengukuran kadar hemoglobin diukur dengan metode Sahli dengan Sahlinometer (Wedemeyer dan Yasutake, 1977).
Tabung sahlinometer disi dengan larutan HCl 0,1 N sampai angka 10 (garis skala paling bawah pada tabung sahlinometer). Tabung tersebut ditempatkan diantara dua tabung dengan warna standar. Darah ikan diambil dari tabung ependorf dengan pipet sahli sebanyak 0,02 mL dan dimasukkan ke dalam tabung sahli dan diamkan selama 3 menit. Akuades ditambahkan ke dalam tabung dengan pipet tetes sedikit demi sedikit sambil diaduk dengan gelas pengaduk sampai warnanya tepat sama dengan warna standar. Pembacaan skala dilakukan dengan melihat permukaan cairan dan dicocokkan dengan skala tabung sahli yang dilihat pada skala jalur gr % (kuning) yang berarti banyaknya haemoglobin dalam gram per 100 mL darah. Kadar hemoglobin dinyatakan dalam gr %.
2. Pengukuran hematokrit dengan metode Anderson dan Siwicki (1995)
Sampel darah dimasukkan dalam tabung mikrohematokrit sampai kira-kira 4/5 bagian tabung, sumbat ujungnya (berwarna merah) dengan kretoseal. Hematokrit disentrifus dengan kecepatan 3500 rpm selama 15 menit. Panjang bagian darah yang mengendap (a), panjang total volume darah dalam tabung (b) diukur.
Kadar hematokrit (He) = (a/b) x 100% ;
kadar He dinyatakan sebagai % volume padatan sel darah. 3. Pengukuran total leukosit dengan metode Blaxhall dan Daisley (1973)
Sampel darah dihisap dari eppendorf dengan pipet sampai skala 0,5 (pipet yang digunakan adalah pipet khusus pengukuran leukosit), dilanjutkan dengan
menghisap larutan Turk’s sampai skala 11, goyangkan pipet agar bercampur
homogen. Tetesan pertama dibuang, tetesan berikutnya dimasukkan ke dalam hemasitometer dan tutup dengan kaca penutup. Penghitungan dilakukan pada 5 kotak besar hemasitometer. Jumlah leukosit dihitung dengan cara mengkonversikan dengan jumlah total kotak besar dan volume kotak sehingga didapat jumlah sel darah putih per mm3.
4. Pengukuran diferensial leukosit dengan metode Blaxhall dan Daisley (1973) Membuat sediaan ulas darah, keringkan di udara dan difiksasi dengan methanol 5 menit, lalu dikeringkan di udara. Selanjutnya diwarnai dengan cara merendam ke dalam pewarna Giemsa (7%) selama 15 menit, kemudian dicuci dengan air mengalir dan dikeringkan di atas kertas tissu. Jenis-jenis leukosit dihitung sampai berjumlah 100 sel.
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5. Pengukuran total eritrosit dengan metode Blaxhall dan Daisley (1973)
Sampel darah dihisap dengan pipet berskala sampai 0,5, selanjutnya menghisap larutan Hayem sampai skala 101, goyangkan agar bercampur homogen. Tetesan pertama dibuang, berikutnya diteteskan dalam hemasitometer dan tutup dengan kaca penutup,
Penghitungan dilakukan pada 5 kotak kecil hemasitometer, Jumlah eritrosit = rataan Σ sel terhitung x 1
0.2�0.2�0.1 x pengenceran (sel/mm3)
6. Pengukuran aktivitas fagositosis, metode Anderson dan Siwicki (1995); Watanuki et al. (2006)
Mengambil sebanyak 50 µl darah, dimasukkan ke dalam tabung ependorf, ditambahkan 50 µl suspensi Staphylococcus aerus selama 20 menit. Membuat sediaan ulas dan dikering udarakan, difiksasi dengan ethyl alkohol (95%) selama 5 menit, dikeringkan, selanjutnya diwarnai dengan cara merendam