Detection of leukemia markers using long-range surface plasmon waveguides functionalized with Protein G

A novel optical biosensor based on long-range surface plasmon-polariton (LRSPP) waveguides is demonstrated for the detection of leukemia markers in patient serum using a functionalization strategy based on Protein G. The sensor consists of thin straight Au waveguides (5 mu m x 35 nm x 3.2 mm) embedded in fluoropolymer CYTOP (TM) with a fluidic channel etched into the top cladding. B-cell leukemia is characterized by a high B-cell count and abnormal distribution of immunoglobulin G kappa (IgG kappa) and lambda (IgG lambda) light chains in serum. The detection of leukemic abnormalities in serum was performed based on determining IgG kappa-to-IgG lambda ratios (kappa : lambda). Three patient sera were tested: high kappa (HKS, kappa : lambda similar to 12.7 : 1), high lambda (HLS, lambda : kappa similar to 6.9 : 1) and normal (control) sera (NS, kappa : lambda similar to 1.7:1). Au waveguides were functionalized with Protein G and two complementary immobilization approaches were investigated: a) the reverse approach, where the Protein G surface is functionalized with patient serum and then tested against goat anti-human IgG light chains in buffer, and b) the direct approach, where the Protein G surface is functionalized with goat anti-human IgGs first and then tested against patient serum. The reverse approach was found to be more effective and robust because Protein G-functionalized surface performs as an immunological filter by capturing primarily IgGs out of the pool of serum proteins. For the reverse approach, the ratios measured were 3.7 : 1(kappa : lambda), 9.7 : 1(lambda : kappa) and 1.9 : 1(kappa : lambda) for HKS, HLS and NS, respectively, which compare favorably with corresponding protein densitometry measurements. The respective ratios for the direct approach were 2.6 : 1(kappa : lambda), 2.6 : 1(lambda : kappa) and 1.7 : 1(kappa : lambda). The binding strength and cross-reactivity of goat anti-human IgGs light chains were also determined using pure solutions. The LRSPP biosensor along with the innovative reverse approach can provide a low-cost and compact solution to B-cell leukemia screening.