Comparison of sheep erythrocytes and Korean native goat erythrocytes-rosette forming rate of pig peripheral blood mononuclear cells

돼지 말초혈액 단핵세포의 면양 및 재래산양 적혈구 rosette 형성능 비교

To develope the methods for isolation and enumeration of lymphocyte subpopulations in pigs, we carried out the rosette-assay using sheep erythrocytes(SRBC) and Korean native goat erythrocytes(GRBC) as a target cells. To enumerate T lymphocytes, E-rosette methods were applied with RBC treated with various concentration of polymers such as Aet and Dex, singly or in combination. And to enumerate B lymphocytes, EAand EAC-rosette assay was adopted. The results were as follows; 1. E-RFR with polymer-untreated SRBC and GRBC was $32.9{\pm}7.9%$ and $31.3{\pm}9.4%$, respectively. On the other hand, RFR with 0.1M Aet plus 8% Dex treated SRBC and GRBC was increased about two-fold($67.8{\pm}7.4%$ and $69.8{\pm}8.5%$), respectively. 2. EA-RFR with SRBC and GRBC were $ 39.1{\pm}10.2%$ and $32.6{\pm}6.1%$, respectively. 3. EAC-RFR with SRBC and GRBC were $27.6{\pm}7.0%$ arld $21.0{\pm}3.2%$, respectively. These results showed that both SRBC and GRBC could be recommanded as a binding cells for rosetteassay to isolate of lymphocyte-subpopulations in pigs.

돼지(n=22) 순환혈액내 단핵세포(PB-MNCs)중 T 및 B 림프구를 정량하고자 rosette 형성기법을 적용하였다. T 림프구는 여러 농도의 2-aminoethylisothiouronium bromide(Aet)나 dextran(Dex) 용액 또는 Aet와 Dex로 조합 처리된 면양(SRBC) 및 재래산양(GRBC)의 적혈구를 사용한 E-rosette 법으로, B 림프구는 erythrocyeantibody(EA) 및 erythrocyte-antibody-complement(EAC) rosette 법으로 각각 정량하였다. 한편 rosette 형성세포의 냉장정치시간에 따른 판독결과를 비교하여 적정판독시간대를 구명한 바 아래의 결과를 얻었다. 1. PB-MNCs와 SRBC/GRBC와의 자연 rosette형성을(RFR)은$ 32.9{\pm}7.9%/31.3{\pm}9.4%$인데 비하여 Aet 또는 Dex 처리군에서는 모두 증가되었고 특히 0.15M Aet$(40.2{\pm}4.6%/37.0{\pm}3.3%)$ 및 8% Dex$(71.5{\pm}4.5%/69.9{\pm}5.8%)$ 처리군에서 각각 높게 나타났다. 한편 0.1M Aet처리후 8% Dex 첨가군에서는 $67.8{\pm}7.4%$ 및 $69.8{\pm}8.5%$로 나타나 복잡한 Aet와 Dex를 복합처리하기 보다는 8% Dex 단독처리가 간편함을 알 수 있었다. 2. Rosette 형성세포의 냉장보관중 최적판독시간대는 10~20시간의 범위이었다. 3. EA 및 EAC의 RFR은 SRBC의 경우 $39.1{\pm}10.2%$, $27.6{\pm}7.0%$, GRBC의 경우 $32.6{\pm}6.1%$, $21.0{\pm}3.2%$로 나타났다. 이러한 결과는 돼지 PB-MNCs내의 T 및 B 림프구를 rosette 형성법으로 분리할 수 있으며 rosette assay에서 SRBC 뿐아니라 GRBC의 이용이 가능함을 제시한다.