Assay for detection of antibodies to lysosomal enzymes
View Patent ↗The present invention relates to methods for screening body fluids or tissues for antibodies, including neutralizing and isotype-specific antibodies, against lysosomal enzymes administered as part of enzyme replacement therapy.
1. A method for detecting lysosomal enzyme (LE)-specific antibodies in a human comprising the steps of:
(a) contacting a body fluid sample from the human with a first lysosomal enzyme labeled with a capture moiety and a second lysosomal enzyme labeled with a detection moiety to form a LE-specific antibody/lysosomal enzyme complex,
(b) capturing the LE-specific antibody/lysosomal enzyme from step (a) by contacting the LE-specific antibody/lysosomal enzyme complex with a solid support that specifically binds to the capture moiety, and
(c) detecting the presence of captured LE-specific antibody from the body fluid sample by detecting the presence of the second lysosomal enzyme labeled with the detection moiety.
2. The method of claim 1 , wherein the capture moiety of the first lysosomal enzyme is bound to the solid support via non-covalent interaction.
3. The method of claim 2 , wherein the non-covalent interaction is biotin-avidin interaction.
4. The method of claim 1 , wherein the body fluid, first lysosomal enzyme and second lysosomal enzyme are all contacted together in the same solution.
5. The method of claim 1 , wherein the body fluid, first lysosomal enzyme and second lysosomal enzyme are contacted together for at least about 30 minutes prior to step (b).
6. The method of claim 1 , wherein the molar concentration of the first lysosomal enzyme is about the same or less than the molar concentration of the second lysosomal enzyme.
7. The method of claim 1 , wherein the first lysosomal enzyme and the second lysosomal enzyme are used in about equimolar ratios.
8. The method of claim 1 , wherein the limit of detection is less than about 500 ng/mL.
9. The method of claim 1 , wherein the limit of detection is less than about 100 ng/mL.
10. The method of claim 1 , wherein the limit of detection is about 1.7 ng/mL to about 8.5 ng/mL.
11. The method of claim 1 , wherein LE-specific antibodies can be detected from a species selected from the group consisting of human, cynomolgus monkey, feline, canine, rabbit, rat and mouse.
12. The method of claim 11 , wherein the species is human.
13. The method of claim 1 , wherein the body fluid is serum.
14. The method of claim 1 , wherein the first and second lysosomal enzyme are recombinant human arylsulfatase (rhASB).
15. The method of claim 1 , wherein the first and second lysosomal enzyme are recombinant human α-glucosidase (rhGAA).
16. A method for detecting lysosomal enzyme (LE)-specific antibodies in a human comprising the steps of:
(a) capturing the LE-specific antibody in a body fluid sample from the human by contacting the body fluid sample with a first lysosomal enzyme bound to a solid support,
(b) contacting captured LE-specific antibody from step (a) with a second lysosomal enzyme labeled with a detection moiety, and
(c) detecting the presence of captured LE-specific antibody from the serum sample by detecting the presence of the second lysosomal enzyme labeled with the detection moiety.